Disease resistant plant methods and compositions

ABSTRACT

The present invention provides methods and compositions for producing elite lines of corn exhibiting anthracnose stalk rot (ASR) resistance. Also provided in the present invention are corn plants exhibiting ASR resistance resulting from such methods, and methods for breeding corn such that the ASR resistance traits may be transferred to a desired genetic background.

REFERENCE TO RELATED APPLICATIONS

This application is a divisional of U.S. application Ser. No.16/831,282, filed Mar. 26, 2020, is a divisional of U.S. applicationSer. No. 16/404,570, filed May 6, 2019, now U.S. Pat. No. 10,638,685,which is a divisional of U.S. application Ser. No. 14/801,618, filedJul. 16, 2015, now U.S. Pat. No. 10,280,433, which claims the benefit ofU.S. Provisional Application No. 62/027,153, filed Jul. 21, 2014, andU.S. Provisional Application No. 62/101,292, filed Jan. 8, 2015 each ofwhich is herein incorporated by reference in their entireties.

FIELD OF THE INVENTION

The present invention relates to the field of agriculturalbiotechnology. More specifically, the invention relates to methods forproducing corn plants with resistance to fungi.

INCORPORATION OF SEQUENCE LISTING

A sequence listing contained in the file named “MONS358US_ST25.txt”which is 40 bytes (measured in MS-Windows®) and created on Jul. 16,2015, and comprises 106 nucleotide sequences, is filed electronicallyherewith and incorporated by reference in its entirety.

BACKGROUND OF THE INVENTION

Advances in molecular genetics have made it possible to select plantsbased on genetic markers linked to traits of interest, a process calledmarker-assisted selection (MAS). While breeding efforts to date haveprovided a number of useful corn lines and varieties with beneficialtraits, there remains a need in the art for selection of varieties withfurther improved traits and methods for their production. In many cases,such efforts have been hampered by difficulties in identifying and usingalleles conferring beneficial traits. These efforts can be confounded bythe lack of definitive phenotypic assays, and other issues such asepistasis and polygenic or quantitative inheritance. In the absence ofmolecular tools such as MAS, it may not be practical to attempt toproduce certain new genotypes of crop plants due to such challenges.

SUMMARY OF THE INVENTION

In one aspect, the present invention provides a method for obtaining acorn plant with enhanced anthracnose stalk rot resistance comprising: a)providing a population of corn plants; b) detecting in said plants ananthracnose stalk rot resistance allele at a polymorphic locus in achromosomal segment flanked by loci IDP7601 and gpm426b on chromosome 6;c) selecting from said population at least a first plant comprising saidallele and enhanced anthracnose stalk rot resistance compared to a plantlacking said allele. In some embodiments, said segment is flanked bymarker loci umc2006 and chs562. In other embodiments, said segment isflanked by marker loci umc2006 and SEQ ID NO: 8. In further embodiments,said segment is flanked by marker loci SEQ ID NO: 52 and SEQ ID NO: 8.In yet further embodiments, said segment is flanked by marker loci SEQID NO: 4 and SEQ ID NO: 2. In other embodiments, said segment is flankedby marker loci SEQ ID NO: 96 and SEQ ID NO: 106. In some embodiments,said polymorphic locus is selected from the group consisting of:IDP7601, IDP62, 111, IDP8090, umc2006, IDP8231, umc248b, SEQ ID NO: 10,pco136292, SEQ ID NO: 3, IDP6025, IDP6010, SEQ ID NO: 1, SEQ ID NO: 7,agrr118a, umc180 (pep), SEQ ID NO: 51, gpm74, TIDP3136, AY107053, SEQ IDNO: 4, SEQ ID NO: 52, IDP1699, pdi7, gpm869, SEQ ID NO: 81, ufg11, SEQID NO: 82, umc1250, SEQ ID NO: 53, TIDP3356, SEQ ID NO: 83, SEQ ID NO:96, csu382a (cld), IDP2409, SEQ ID NO: 97, SEQ ID NO: 98, SEQ ID NO: 99,SEQ ID NO: 100, SEQ ID NO: 101, SEQ ID NO: 102, SEQ ID NO: 103, SEQ IDNO: 104, SEQ ID NO: 105, SEQ ID NO: 106, PCO146525, SEQ ID NO: 8,csu225, bnl3.03, AI665560, SEQ ID NO: 54, pzb00414, umc2141, SEQ ID NO:55, SEQ ID NO: 2, AY110435, elfa5, SEQ ID NO: 5, umc1379, bnl15.37a, SEQID NO: 56, pza02478, IDP3886, c139957_1, mmc0241, dup400 (pac),jpsb107b, chs562, gpm709b, SEQ ID NO: 6, umc2321, bnlg1702, SEQ ID NO:9, csu158b (eno), and gpm426b. In further embodiments, the inventionprovides a corn plant produced by the methods provided herein, or aplant part or seed of said corn plant.

In another aspect, the present invention provides a method of producinga corn plant with enhanced anthracnose stalk rot resistance comprising:a) introgressing into a corn plant a genomic segment comprising ananthracnose stalk rot resistance allele; and b) selecting a plant basedon the presence of said allele in at least one polymorphic locusselected from the group consisting of: IDP7601, IDP62, 111, IDP8090,umc2006, IDP8231, umc248b, SEQ ID NO: 10, pco136292, SEQ ID NO: 3,IDP6025, IDP6010, SEQ ID NO: 1, SEQ ID NO: 7, agrr118a, umc180 (pep),SEQ ID NO: 51, gpm74, TIDP3136, AY107053, SEQ ID NO: 4, SEQ ID NO: 52,IDP1699, pdi7, gpm869, SEQ ID NO: 81, ufg11, SEQ ID NO: 82, umc1250, SEQID NO: 53, TIDP3356, SEQ ID NO: 83, SEQ ID NO: 96, csu382a (cld),IDP2409, SEQ ID NO: 97, SEQ ID NO: 98, SEQ ID NO: 99, SEQ ID NO: 100,SEQ ID NO: 101, SEQ ID NO: 102, SEQ ID NO: 103, SEQ ID NO: 104, SEQ IDNO: 105, SEQ ID NO: 106, PCO146525, SEQ ID NO: 8, csu225, bnl3.03,AI665560, SEQ ID NO: 54, pzb00414, umc2141, SEQ ID NO: 55, SEQ ID NO: 2,AY110435, elfa5, SEQ ID NO: 5, umc1379, bnl15.37a, SEQ ID NO: 56,pza02478, IDP3886, c139957_1, mmc0241, dup400 (pac), jpsb107b, chs562,gpm709b, SEQ ID NO: 6, umc2321, bnlg1702, SEQ ID NO: 9, csu158b (eno),and gpm426b; wherein said allele confers enhanced resistance toanthracnose stalk rot compared to a plant lacking said allele. Infurther embodiments, the method further comprises: c) crossing said cornplant with itself or a second plant to produce one or more progenyplants; and d) selecting a progeny plant comprising said allele. In someembodiments, step (d) of selecting comprises marker-assisted selection.In other embodiments, the progeny plant is an F2-F6 progeny plant. Infurther embodiments, producing the progeny plant comprises backcrossing.In yet further embodiments, backcrossing comprises from 2-7 generationsof backcrosses. In certain embodiments, backcrossing comprisesmarker-assisted selection in at least two generations. In furtherembodiments, the invention provides a corn plant produced by the methodsprovided herein, or a plant part or seed of said corn plant.

In yet another aspect, the invention provides a method of producing acorn plant with enhanced anthracnose stalk rot resistance comprising: a)crossing a first corn plant comprising an anthracnose stalk rotresistance allele with a second corn plant of a different genotype toproduce one or more progeny plants; and b) selecting a progeny plantbased on the presence of said allele in at least one polymorphic locusselected from the group consisting of: IDP7601, IDP62, 111, IDP8090,umc2006, IDP8231, umc248b, SEQ ID NO: 10, pco136292, SEQ ID NO: 3,IDP6025, IDP6010, SEQ ID NO: 1, SEQ ID NO: 7, agrr118a, umc180 (pep),SEQ ID NO: 51, gpm74, TIDP3136, AY107053, SEQ ID NO: 4, SEQ ID NO: 52,IDP1699, pdi7, gpm869, SEQ ID NO: 81, ufg11, SEQ ID NO: 82, umc1250, SEQID NO: 53, TIDP3356, SEQ ID NO: 83, SEQ ID NO: 96, csu382a (cld),IDP2409, SEQ ID NO: 97, SEQ ID NO: 98, SEQ ID NO: 99, SEQ ID NO: 100,SEQ ID NO: 101, SEQ ID NO: 102, SEQ ID NO: 103, SEQ ID NO: 104, SEQ IDNO: 105, SEQ ID NO: 106, PCO146525, SEQ ID NO: 8, csu225, bnl3.03,AI665560, SEQ ID NO: 54, pzb00414, umc2141, SEQ ID NO: 55, SEQ ID NO: 2,AY110435, elfa5, SEQ ID NO: 5, umc1379, bnl15.37a, SEQ ID NO: 56,pza02478, IDP3886, c139957_1, mmc0241, dup400 (pac), jpsb107b, chs562,gpm709b, SEQ ID NO: 6, umc2321, bnlg1702, SEQ ID NO: 9, csu158b (eno),and gpm426b; wherein said allele confers enhanced resistance toanthracnose stalk rot compared to a plant lacking said allele. In someembodiments, step (b) of selecting comprises marker-assisted selection.In other embodiments, the progeny plant is an F2-F6 progeny plant. Infurther embodiments, producing the progeny plant comprises backcrossing.In yet further embodiments, backcrossing comprises from 2-7 generationsof backcrosses. In some embodiments, backcrossing comprisesmarker-assisted selection in at least two generations. In otherembodiments, the first corn plant is an inbred or a hybrid. In furtherembodiments, the second corn plant is an agronomically elite corn plant.In yet further embodiments, the agronomically elite corn plant is aninbred or a hybrid. In further embodiments, the invention provides acorn plant produced by the methods provided herein, or a plant part orseed of said corn plant.

In some aspects, corn plants or methods disclosed herein are used incombination with one or more pesticides including, but not limited to,herbicides, fungicides, insecticides, microbicides, nematicides, insectrepellents, bactericides, and other substances used to control pests. Inother aspects, the corn plants or methods disclosed herein are used incombination with one or more triazoles, strobilurins, acylamino acids,pyrimidines, pyridines, aryl phenyl ketones, amides, benzanilides,imidazoles, dinitrophenols, morpholines, phenylsulfamides andorganophosphorus cpds, derivatives thereof and combinations thereofwhich may be applied as seed, foliar, drench or drip treatments.

DESCRIPTION OF THE FIGURES

FIG. 1 shows first round fine-mapping of ASR-6.01 from CV820914/CV391950as also shown in Table 10. Bulk “f” shared at least one same allele asthe resistant inbred line (CV391950) at the SNP positions represented bySEQ ID NO: 51, SEQ ID NO: 52 and SEQ ID NO: 53 (highlighted by whitecells). Bulk “f” shared the same alleles as the susceptible inbred line(CV820914) at the SNP positions represented by SEQ ID NO: 8, SEQ ID NO:54, SEQ ID NO: 55, SEQ ID NO: 5 and SEQ ID NO: 56 (highlighted by greycells). Similar experiments were also conducted on BC1F2 kernels derivedfrom CV295879/CV391950. Bulk “f”, “g”, “i”, “j”, “p” and “o” displayedsignificantly reduced ASR severity (highlighted by black box, p-value<=0.05) compared with the bulk “b”. Among these resistant bulks, thesame common SNP (SEQ ID NO: 53, highlighted by black oval) wasidentified as the peak marker.

FIG. 2 shows first round fine-mapping of ASR-6.01 from CV295879/CV391950as also shown in Table 11.

FIG. 3 shows second round fine-mapping of ASR-6.01 fromCV005260/CV391950 as also shown in Table 14. For example, bulk “a”shared the same alleles as the resistant inbred line, CV391950, at thecandidate QTL region (highlighted by white cells); bulk “b” shared thesame alleles as the susceptible inbred line, CV005260, at the candidateQTL region (highlighted by grey cells).

DETAILED DESCRIPTION OF THE INVENTION

Anthracnose stalk rot (ASR) is caused by the fungal pathogenColletotrichum graminicola, and results in severe yield loss in cropplants. Efforts to identify or produce plant lines resistant to ASR havebeen hindered by a limited understanding of the genetic loci controllingASR resistance and a lack of available markers for detecting andtracking ASR resistance in plants. Yield loss due to ASR thereforeremains a significant problem.

The present invention identifies previously-unknown genetic loci whichconfer ASR resistance and provides novel molecular markers linked to ASRresistance in plants. The invention further provides methods forintrogression of genetic loci conferring ASR resistance into plantvarieties previously lacking such loci, thereby providing plants with anew or improved disease resistance. The genetic loci, markers, andmethods provided by the invention therefore represent a significantadvance in the art, enabling production of new varieties exhibiting ASRresistance.

In some embodiments, the invention therefore provides quantitative traitloci (QTL) that demonstrate significant co-segregation with ASRresistance. The QTL of the invention can be tracked during plantbreeding or introgressed into a desired genetic background in order toprovide novel plants exhibiting ASR resistance and one or more otherbeneficial traits. In particular embodiments, the invention identifiesfor the first time a QTL on chromosome 6 of the corn genome, designatedASR-6.01, which is associated with ASR resistance.

In other embodiments, the invention provides molecular markers linked tothe QTL of the invention and methods of using the markers for detectionof and selection for ASR resistance. Embodiments of the inventiontherefore include specific markers, chromosome intervals comprising themarkers, and methods of detecting markers genetically linked to ASR-6.01to identify disease resistant plant lines. For example, the inventionprovides a chromosome interval associated with ASR resistance which isflanked by the markers IDP7601 and gpm426b, and which comprises markershaving SEQ ID NOs: 1-10, 51-56, 81-83, and 96-106, or any of the markerslisted in Table 17, and any other markers genetically linked thereto.Also provided herein are markers that are useful for detecting thepresence or absence of disease resistance alleles within the QTL of theinvention that can be used in marker assisted selection (MAS) breedingprograms to produce plants with improved resistance to ASR infection.

The invention further provides methods of using the markers identifiedherein to introgress loci associated with ASR resistance into plants.Thus, one skilled in the art can use the invention to create novel maizeplants with ASR resistance by crossing a donor line comprising a QTLassociated with ASR resistance into any desired recipient line, with orwithout MAS. Resulting progeny can be selected to be genetically similarto the recipient line except for the ASR resistance QTL.

Quantitative Trait Loci

The term “chromosome interval” designates a contiguous linear span ofgenomic DNA that resides on a single chromosome. A chromosome intervalmay comprise a QTL linked with a genetic trait and the QTL may comprisea single gene or multiple genes associated with the genetic trait. Theboundaries of a chromosome interval comprising a QTL are drawn such thata marker that lies within the chromosome interval can be used as amarker for the genetic trait, as well as markers genetically linkedthereto. Each interval comprising a QTL comprises at least one geneconferring a given trait, however knowledge of how many genes are in aparticular interval is not necessary to make or practice the invention,as such an interval will segregate at meiosis as a linkage block. Inaccordance with the invention, a chromosomal interval comprising a QTLmay therefore be readily introgressed and tracked in a given geneticbackground using the methods and compositions provided herein.

Identification of chromosomal intervals and QTL is therefore beneficialfor detecting and tracking a genetic trait, such as ASR resistance, inplant populations. In some embodiments, this is accomplished byidentification of markers linked to a particular QTL. The principles ofQTL analysis and statistical methods for calculating linkage betweenmarkers and useful QTL include penalized regression analysis, ridgeregression, single point marker analysis, complex pedigree analysis,Bayesian MCMC, identity-by-descent analysis, interval mapping, compositeinterval mapping (CIM), and Haseman-Elston regression. QTL analyses maybe performed with the help of a computer and specialized softwareavailable from a variety of public and commercial sources known to thoseof skill in the art.

In some embodiments, the invention provides a chromosomal intervalcomprising a QTL associated with ASR resistance. The invention providesmultiple markers associated with ASR resistance, for example the markershaving the sequence of SEQ ID NOs: 1-10, 51-56, 81-83, or 96-106. Theinvention therefore provides a plant comprising a nucleic acid moleculeselected from the group consisting of SEQ ID NOs: 1-10, 51-56, 81-83,96-106, fragments thereof, or complements thereof. The present inventionfurther provides a plant comprising alleles of the chromosome intervallinked to ASR resistance or fragments and complements thereof as well asany plant comprising any combination of one or more disease resistanceloci selected from the group consisting of SEQ ID NOs: 1-10, 51-56,81-83 and 96-106. Plants provided by the invention may be homozygous orheterozygous for such alleles.

In one embodiment, the chromosome interval associated with ASRresistance contains SEQ ID NOs: 1-10, 51-56, 81-83 or 96-106, and isflanked by the markers IDP7601 and gpm426b. This chromosome intervalencompasses markers that co-segregate with ASR resistance in thepopulations studied at a p-value ≤0.05. An example of a subintervalassociated with ASR resistance includes the interval flanked by umc2006and chs562, which define a chromosome interval encompassing markers thatco-segregate with ASR resistance in populations studied at a p-level≤0.05. An example of a subinterval associated with ASR resistanceincludes the interval flanked by SEQ ID NO: 52 and SEQ ID NO: 8, whichdefine a chromosome interval encompassing markers that co-segregate withASR resistance in populations studied at a p-level ≤0.05. A furtherexample of a subinterval associated with ASR resistance includes theinterval flanked by SEQ ID NO: 53 and SEQ ID NO: 8, that define achromosome interval encompassing markers that co-segregate with ASRresistance in the populations studied at a p-level ≤0.05. Anotherexample of a subinterval associated with ASR resistance includes theinterval flanked by SEQ ID NO: 96 and SEQ ID NO: 106, that define achromosome interval encompassing markers that co-segregate with ASRresistance in the populations studied at a p-level ≤0.001.

Thus, one skilled in the art can use the invention to create novel maizeplants with ASR resistance by associating disease resistance phenotypeswith genotypes at previously unknown disease resistance loci in themaize genome. Disclosed herein are chromosome intervals that comprisealleles responsible for phenotypic differences between ASR resistant andASR susceptible corn lines. The chromosome intervals of the inventionare characterized in specific embodiments by genomic regions includingand flanked by the markers IDP7601 and gpm426b, which comprise markerswithin or closely linked to (within 20 cM of) ASR-6.01. The inventionalso comprises other intervals whose borders fall between, andincluding, those of IDP7601 and gpm426b, or any interval closely linkedto those intervals.

Examples of markers useful for this purpose comprise the SNP markerslisted in Table 16, or any marker linked thereto, including a markerthat maps within or is genetically linked to the chromosome intervalsdescribed herein, including the termini of the intervals. Such markerscan be assayed simultaneously or sequentially in a single sample orpopulation of samples.

Accordingly, the compositions and methods of the present invention canbe utilized to guide MAS or breeding maize varieties with a desiredcomplement (set) of allelic forms of chromosome intervals associatedwith superior agronomic performance (resistance, along with any otheravailable markers for yield, disease resistance, etc.). Any of thedisclosed marker alleles can be introduced into a corn line viaintrogression, by traditional breeding (or introduced viatransformation, or both) to yield a corn plant with superior agronomicperformance. The number of alleles associated with resistance that canbe introduced or be present in a corn plant of the present inventionranges from 1 to the number of alleles disclosed herein, each integer ofwhich is incorporated herein as if explicitly recited.

MAS using additional markers flanking either side of the DNA locusprovide further efficiency because an unlikely double recombinationevent would be needed to simultaneously break linkage between the locusand both markers. Moreover, using markers tightly flanking a locus, oneskilled in the art of MAS can reduce linkage drag by more accuratelyselecting individuals that have less of the potentially deleteriousdonor parent DNA. Any marker linked to or among the chromosome intervalsdescribed herein can thus find use within the scope of this invention.

Similarly, by identifying plants lacking a desired marker locus,susceptible or less resistant plants can be identified, and eliminatedfrom subsequent crosses. Similarly, these marker loci can beintrogressed into any desired genomic background, germplasm, plant,line, variety, etc., as part of an overall MAS breeding program designedto enhance disease resistance. The invention also provides chromosomeQTL intervals that find use in MAS to select plants that demonstratedisease resistance or improved tolerance. The QTL intervals can also beused to counter-select plants that are susceptible or have reducedresistance to disease.

The present invention also extends to a method of making a progeny cornplant and the resulting progeny corn plants. The method comprises, in anembodiment, crossing a first parent corn plant with a second corn plantand growing the female corn plant parent under plant growth conditionsto yield corn plant progeny. Methods of crossing and growing corn plantsare well within the ability of those of ordinary skill in the art. Suchcorn plant progeny can be assayed for alleles associated with ASRresistance as disclosed herein and, thereby, the desired progenyselected. Such progeny plants or seed thereof can be sold commerciallyfor corn production, used for food, processed to obtain a desiredconstituent of the corn, or further utilized in subsequent rounds ofbreeding. At least one of the first or second corn plants is a cornplant of the present invention in that it comprises at least one of theallelic forms of the markers of the present invention, such that theprogeny are capable of inheriting the allele.

Often, a method of the present invention may be applied to at least onerelated corn plant such as from progenitor or descendant line in thesubject corn plants' pedigree such that inheritance of the desiredresistance allele can be traced. The number of generations separatingthe corn plants being subjected to the methods of the present inventionmay be, in specific embodiments, from 1 to 20, commonly 1 to 5, andincluding 1, 2, or 3 generations of separation, and often a directdescendant or parent of the corn plant will be subject to the method(i.e., one generation of separation).

Thus, the invention permits one skilled in the art to detect thepresence or absence of disease resistance genotypes in the genomes ofcorn plants as part of a MAS program. In one embodiment, a breederascertains the genotype at one or more markers for a disease resistantparent, which contains a disease resistance allele, and the genotype atone or more markers for a susceptible parent, which lacks the resistanceallele. For example, the markers of the present invention can be used inMAS in crosses involving elite x exotic corn lines by subjecting thesegregating progeny to MAS to maintain disease resistance alleles, oralleles associated with yield under disease conditions. A breeder canthen reliably track the inheritance of the resistance alleles throughsubsequent populations derived from crosses between the two parents bygenotyping offspring with the markers used on the parents and comparingthe genotypes at those markers with those of the parents. Depending onhow tightly linked the marker alleles are with the trait, progeny thatshare genotypes with the disease resistant parent can be reliablypredicted to express the resistant phenotype and progeny that sharegenotypes with the disease susceptible parent can be reliably predictedto express the susceptible phenotype. Thus, the laborious, inefficient,and potentially inaccurate process of manually phenotyping the progenyfor disease resistance is avoided.

By providing the positions in the maize genome of the intervals and thedisease resistance associated markers within, this invention also allowsone skilled in the art to identify and use other markers within theintervals disclosed herein or linked to the chromosome intervalsdisclosed herein. Having identified such regions, these markers can bereadily identified from public linkage maps.

Closely linked markers flanking the locus of interest that have allelesin linkage disequilibrium with a resistance allele at that locus may beeffectively used to select for progeny plants with enhanced resistanceto disease conditions. Thus, the markers described herein, such as thoselisted in Table 16, as well as other markers genetically linked to thesame chromosome interval, may be used to select for maize plants withenhanced resistance to ASR. Often, a set of these markers will be used,(e.g., 2 or more, 3 or more, 4 or more, 5 or more) in the flankingregions of the gene. Optionally, as described above, a marker flankingon either side or within the actual gene and/or locus may also be used.The parents and their progeny may be screened for these sets of markers,and the markers that are polymorphic between the two parents used forselection. In an introgression program, this allows for selection of thegene or locus genotype at the more proximal polymorphic markers andselection for the recurrent parent genotype at the more distalpolymorphic markers.

The choice of markers actually used to practice the invention is notlimited and can be any marker that is genetically linked to theintervals described herein, which includes markers mapping within theintervals. One example includes any marker selected from SEQ ID NOs:1-10, 51-56, 81-83, 96-106, or the markers listed in Table 17.Furthermore, since there are many different types of marker detectionassays known in the art, it is not intended that the type of markerdetection assay used to practice this invention be limited in any way.

Molecular Markers

“Marker,” “genetic marker,” “molecular marker,” “marker nucleic acid,”and “marker locus” refer to a nucleotide sequence or encoded productthereof (e.g., a protein) used as a point of reference when identifyinga linked locus. A marker can be derived from genomic nucleotide sequenceor from expressed nucleotide sequences (e.g., from a spliced RNA, acDNA, etc.), or from an encoded polypeptide, and can be represented byone or more particular variant sequences, or by a consensus sequence. Inanother sense, a marker is an isolated variant or consensus of such asequence. The term also refers to nucleic acid sequences complementaryto or flanking the marker sequences, such as nucleic acids used asprobes or primer pairs capable of amplifying the marker sequence. A“marker probe” is a nucleic acid sequence or molecule that can be usedto identify the presence of a marker locus, e.g., a nucleic acid probethat is complementary to a marker locus sequence. Alternatively, in someaspects, a marker probe refers to a probe of any type that is able todistinguish (i.e., genotype) the particular allele that is present at amarker locus. A “marker locus” is a locus that can be used to track thepresence of a second linked locus, e.g., a linked locus that encodes orcontributes to expression of a phenotypic trait. For example, a markerlocus can be used to monitor segregation of alleles at a locus, such asa QTL, that are genetically or physically linked to the marker locus.Thus, a “marker allele,” alternatively an “allele of a marker locus” isone of a plurality of polymorphic nucleotide sequences found at a markerlocus in a population that is polymorphic for the marker locus.

“Marker” also refers to nucleic acid sequences complementary to thegenomic sequences, such as nucleic acids used as probes. Markerscorresponding to genetic polymorphisms between members of a populationcan be detected by methods well-established in the art. These include,e.g., PCR-based sequence specific amplification methods, detection ofrestriction fragment length polymorphisms (RFLP), detection of isozymemarkers, detection of polynucleotide polymorphisms by allele specifichybridization (ASH), detection of amplified variable sequences of theplant genome, detection of self-sustained sequence replication,detection of simple sequence repeats (SSRs), detection of singlenucleotide polymorphisms (SNPs), or detection of amplified fragmentlength polymorphisms (AFLPs). Well established methods are also know forthe detection of expressed sequence tags (ESTs) and SSR markers derivedfrom EST sequences and randomly amplified polymorphic DNA (RAPD).

A favorable allele of a marker is the allele of the marker thatco-segregates with a desired phenotype (e.g., disease resistance). Asused herein, a QTL marker has a minimum of one favorable allele,although it is possible that the marker might have two or more favorablealleles found in the population. Any favorable allele of that marker canbe used advantageously for the identification and construction ofdisease resistant plant lines. Optionally, one, two, three or morefavorable allele(s) of different markers are identified in, orintrogressed into a plant, and can be selected for or against duringMAS. Desirably, plants or germplasm are identified that have at leastone such favorable allele that positively correlates with diseaseresistance or improved disease resistance. Alternatively, a markerallele that co-segregates with disease susceptibility also finds usewith the invention, since that allele can be used to identify andcounter select disease susceptible plants. Such an allele can be usedfor exclusionary purposes during breeding to identify alleles thatnegatively correlate with resistance, to eliminate susceptible plants orgermplasm from subsequent rounds of breeding.

The more tightly linked a marker is with a DNA locus influencing aphenotype, the more reliable the marker is in MAS, as the likelihood ofa recombination event unlinking the marker and the locus decreases.Markers containing the causal mutation for a trait, or that are withinthe coding sequence of a causative gene, are ideal as no recombinationis expected between them and the sequence of DNA responsible for thephenotype.

Genetic markers are distinguishable from each other (as well as from theplurality of alleles of any one particular marker) on the basis ofpolynucleotide length and/or sequence. A large number of corn molecularmarkers are known in the art, and are published or available fromvarious sources, such as the MaizeGDB internet resource. In general, anydifferentially inherited polymorphic trait (including a nucleic acidpolymorphism) that segregates among progeny is a potential geneticmarker.

In some embodiments of the invention, one or more marker alleles areselected for in a single plant or a population of plants. In thesemethods, plants are selected that contain favorable alleles from morethan one resistance marker, or alternatively, favorable alleles frommore than one resistance marker are introgressed into a desiredgermplasm. One of skill recognizes that the identification of favorablemarker alleles is germplasm-specific. The determination of which markeralleles correlate with resistance (or susceptibility) is determined forthe particular germplasm under study. One of skill recognizes thatmethods for identifying the favorable alleles are routine and well knownin the art, and furthermore, that the identification and use of suchfavorable alleles is well within the scope of this invention.Furthermore still, identification of favorable marker alleles in plantpopulations other than the populations used or described herein is wellwithin the scope of this invention.

Marker Detection

In some aspects, methods of the invention utilize an amplification stepto detect/genotype a marker locus, but amplification is not always arequirement for marker detection (e.g. Southern blotting and RFLPdetection). Separate detection probes can also be omitted inamplification/detection methods, e.g., by performing a real timeamplification reaction that detects product formation by modification ofthe relevant amplification primer upon incorporation into a product,incorporation of labeled nucleotides into an amplicon, or by monitoringchanges in molecular rotation properties of amplicons as compared tounamplified precursors (e.g., by fluorescence polarization).

“Amplifying,” in the context of nucleic acid amplification, is anyprocess whereby additional copies of a selected nucleic acid (or atranscribed form thereof) are produced. In some embodiments, anamplification-based marker technology is used wherein a primer oramplification primer pair is admixed with genomic nucleic acid isolatedfrom the first plant or germplasm, and wherein the primer or primer pairis complementary or partially complementary to at least a portion of themarker locus, and is capable of initiating DNA polymerization by a DNApolymerase using the plant genomic nucleic acid as a template. Theprimer or primer pair is extended in a DNA polymerization reactionhaving a DNA polymerase and a template genomic nucleic acid to generateat least one amplicon. In other embodiments, plant RNA is the templatefor the amplification reaction. In some embodiments, the QTL marker is aSNP type marker, and the detected allele is a SNP allele, and the methodof detection is allele specific hybridization (ASH).

In general, the majority of genetic markers rely on one or moreproperties of nucleic acids for their detection. Typical amplificationmethods include various polymerase based replication methods, includingthe polymerase chain reaction (PCR), ligase mediated methods such as theligase chain reaction (LCR) and RNA polymerase based amplification(e.g., by transcription) methods. An “amplicon” is an amplified nucleicacid, e.g., a nucleic acid that is produced by amplifying a templatenucleic acid by any available amplification method (e.g., PCR, LCR,transcription, or the like). A “genomic nucleic acid” is a nucleic acidthat corresponds in sequence to a heritable nucleic acid in a cell.Common examples include nuclear genomic DNA and amplicons thereof. Agenomic nucleic acid is, in some cases, different from a spliced RNA, ora corresponding cDNA, in that the spliced RNA or cDNA is processed,e.g., by the splicing machinery, to remove introns. Genomic nucleicacids optionally comprise non-transcribed (e.g., chromosome structuralsequences, promoter regions, enhancer regions, etc.) and/ornon-translated sequences (e.g., introns), whereas spliced RNA/cDNAtypically do not have non-transcribed sequences or introns. A “templatenucleic acid” is a nucleic acid that serves as a template in anamplification reaction (e.g., a polymerase based amplification reactionsuch as PCR, a ligase mediated amplification reaction such as LCR, atranscription reaction, or the like). A template nucleic acid can begenomic in origin, or alternatively, can be derived from expressedsequences, e.g., a cDNA or an EST. Details regarding the use of theseand other amplification methods can be found in any of a variety ofstandard texts. Many available biology texts also have extendeddiscussions regarding PCR and related amplification methods and one ofskill will appreciate that essentially any RNA can be converted into adouble stranded DNA suitable for restriction digestion, PCR expansionand sequencing using reverse transcriptase and a polymerase.

PCR detection and quantification using dual-labeled fluorogenicoligonucleotide probes, commonly referred to as “TaqMan™” probes, canalso be performed according to the present invention. These probes arecomposed of short (e.g., 20-25 base) oligodeoxynucleotides that arelabeled with two different fluorescent dyes. On the 5′ terminus of eachprobe is a reporter dye, and on the 3′ terminus of each probe aquenching dye is found. The oligonucleotide probe sequence iscomplementary to an internal target sequence present in a PCR amplicon.When the probe is intact, energy transfer occurs between the twofluorophores and emission from the reporter is quenched by the quencherby FRET. During the extension phase of PCR, the probe is cleaved by 5′nuclease activity of the polymerase used in the reaction, therebyreleasing the reporter from the oligonucleotide-quencher and producingan increase in reporter emission intensity. TaqMan™ probes areoligonucleotides that have a label and a quencher, where the label isreleased during amplification by the exonuclease action of thepolymerase used in amplification, providing a real time measure ofamplification during synthesis. A variety of TaqMan™ reagents arecommercially available, e.g., from Applied Biosystems as well as from avariety of specialty vendors such as Biosearch Technologies.

In one embodiment, the presence or absence of a molecular marker isdetermined simply through nucleotide sequencing of the polymorphicmarker region. This method is readily adapted to high throughputanalysis as are the other methods noted above, e.g., using availablehigh throughput sequencing methods such as sequencing by hybridization.

In alternative embodiments, in silico methods can be used to detect themarker loci of interest. For example, the sequence of a nucleic acidcomprising the marker locus of interest can be stored in a computer. Thedesired marker locus sequence or its homolog can be identified using anappropriate nucleic acid search algorithm as provided by, for example,in such readily available programs as BLAST, or even simple wordprocessors.

While the exemplary markers provided in the figures and tables hereinare either SNP markers, any of the aforementioned marker types can beemployed in the context of the invention to identify chromosomeintervals encompassing genetic element that contribute to superioragronomic performance (e.g., disease resistance or improved diseasetolerance).

Probes and Primers

In general, synthetic methods for making oligonucleotides, includingprobes, primers, molecular beacons, PNAs, LNAs (locked nucleic acids),etc., are well known. For example, oligonucleotides can be synthesizedchemically according to the solid phase phosphoramidite triester methoddescribed. Oligonucleotides, including modified oligonucleotides, canalso be ordered from a variety of commercial sources.

Nucleic acid probes to the marker loci can be cloned and/or synthesized.Any suitable label can be used with a probe of the invention. Detectablelabels suitable for use with nucleic acid probes include, for example,any composition detectable by spectroscopic, radioisotopic,photochemical, biochemical, immunochemical, electrical, optical orchemical means. Useful labels include biotin for staining with labeledstreptavidin conjugate, magnetic beads, fluorescent dyes, radio labels,enzymes, and colorimetric labels. Other labels include ligands whichbind to antibodies labeled with fluorophores, chemiluminescent agents,and enzymes. A probe can also constitute radio labeled PCR primers thatare used to generate a radio labeled amplicon. It is not intended thatthe nucleic acid probes of the invention be limited to any particularsize.

In some embodiments, the molecular markers of the invention are detectedusing a suitable PCR-based detection method, where the size or sequenceof the PCR amplicon is indicative of the absence or presence of themarker (e.g., a particular marker allele). In these types of methods,PCR primers are hybridized to the conserved regions flanking thepolymorphic marker region. As used in the art, PCR primers used toamplify a molecular marker are sometimes termed “PCR markers” or simply“markers.” It will be appreciated that, although many specific examplesof primers are provided herein, suitable primers to be used with theinvention can be designed using any suitable method. It is not intendedthat the invention be limited to any particular primer or primer pair.In some embodiments, the primers of the invention are radiolabelled, orlabeled by any suitable means (e.g., using a non-radioactive fluorescenttag), to allow for rapid visualization of the different size ampliconsfollowing an amplification reaction without any additional labeling stepor visualization step. In some embodiments, the primers are not labeled,and the amplicons are visualized following their size resolution, e.g.,following agarose gel electrophoresis. In some embodiments, ethidiumbromide staining of the PCR amplicons following size resolution allowsvisualization of the different size amplicons. It is not intended thatthe primers of the invention be limited to generating an amplicon of anyparticular size. For example, the primers used to amplify the markerloci and alleles herein are not limited to amplifying the entire regionof the relevant locus. The primers can generate an amplicon of anysuitable length that is longer or shorter than those disclosed herein.In some embodiments, marker amplification produces an amplicon at least20 nucleotides in length, or alternatively, at least 50 nucleotides inlength, or alternatively, at least 100 nucleotides in length, oralternatively, at least 200 nucleotides in length. Marker alleles inaddition to those recited herein also find use with the presentinvention.

Linkage Analysis

“Linkage”, or “genetic linkage,” is used to describe the degree withwhich one marker locus is associated with another marker locus or someother locus (for example, a resistance locus). A marker locus may belocated within a locus to which it is genetically linked. For example,if locus A has genes “A” or “a” and locus B has genes “B” or “b” and across between parent 1 with AABB and parent 2 with aabb will producefour possible gametes where the genes are segregated into AB, Ab, aB andab. The null expectation is that there will be independent equalsegregation into each of the four possible genotypes, i.e. with nolinkage ¼ of the gametes will of each genotype. Segregation of gametesinto a genotypes differing from ¼ is attributed to linkage. As usedherein, linkage can be between two markers, or alternatively between amarker and a phenotype. A marker locus may be genetically linked to atrait, and in some cases a marker locus genetically linked to a trait islocated within the allele conferring the trait. A marker may also becausative for a trait or phenotype, for example a causativepolymorphism. In a further example, a marker locus can be associatedwith resistance or improved tolerance to a plant pathogen when themarker locus is in linkage disequilibrium with the resistance trait. Thedegree of linkage of a molecular marker to a phenotypic trait (e.g., aQTL) is measured, e.g., as a statistical probability of co-segregationof that molecular marker with the phenotype.

As used herein, “closely linked” means that the marker or locus iswithin about 20 cM, for instance within about 10 cM, about 5 cM, about 1cM, about 0.5 cM, or less than 0.5 cM of the identified locus associatedwith ASR resistance.

As used herein, the linkage relationship between a molecular marker anda phenotype is given is the statistical likelihood that the particularcombination of a phenotype and the presence or absence of a particularmarker allele is random. Thus, the lower the probability score, thegreater the likelihood that a phenotype and a particular marker willcosegregate. In some embodiments, a probability score of 0.05 (p=0.05,or a 5% probability) of random assortment is considered a significantindication of co-segregation. However, the present invention is notlimited to this particular standard, and an acceptable probability canbe any probability of less than 50% (p<0.5). For example, a significantprobability can be less than 0.25, less than 0.20, less than 0.15, orless than 0.1. The phrase “closely linked,” in the present application,means that recombination between two linked loci occurs with a frequencyof equal to or less than about 10% (i.e., are separated on a genetic mapby not more than 10 cM). In one aspect, any marker of the invention islinked (genetically and physically) to any other marker that is at orless than 50 cM distant. In another aspect, any marker of the inventionis closely linked (genetically and physically) to any other marker thatis in close proximity, e.g., at or less than 10 cM distant. Two closelylinked markers on the same chromosome can be positioned 9, 8, 7, 6, 5,4, 3, 2, 1, 0.75, 0.5 or 0.25 cM or less from each other.

Classical linkage analysis can be thought of as a statisticaldescription of the relative frequencies of cosegregation of differenttraits. Linkage analysis is the well characterized descriptive frameworkof how traits are grouped together based upon the frequency with whichthey segregate together. That is, if two non-allelic traits areinherited together with a greater than random frequency, they are saidto be “linked.” The frequency with which the traits are inheritedtogether is the primary measure of how tightly the traits are linked,i.e., traits which are inherited together with a higher frequency aremore closely linked than traits which are inherited together with lower(but still above random) frequency. The further apart on a chromosomethe genes reside, the less likely they are to segregate together,because homologous chromosomes recombine during meiosis. Thus, thefurther apart on a chromosome the genes reside, the more likely it isthat there will be a crossing over event during meiosis that will resultin the marker and the DNA sequence responsible for the trait the markeris designed to track segregating separately into progeny. A commonmeasure of linkage is the frequency with which traits cosegregate.

Linkage analysis is used to determine which polymorphic marker alleledemonstrates a statistical likelihood of co-segregation with theresistance phenotype (thus, a “resistance marker allele”). Followingidentification of a marker allele for co-segregation with the resistancephenotype, it is possible to use this marker for rapid, accuratescreening of plant lines for the resistance allele without the need togrow the plants through their life cycle and await phenotypicevaluations, and furthermore, permits genetic selection for theparticular resistance allele even when the molecular identity of theactual resistance QTL is unknown. Tissue samples can be taken, forexample, from the endosperm, embryo, or mature/developing plant andscreened with the appropriate molecular marker to rapidly determinedetermined which progeny contain the desired genetics. Linked markersalso remove the impact of environmental factors that can often influencephenotypic expression.

Because chromosomal distance is approximately proportional to thefrequency of crossing over events between traits, there is anapproximate physical distance that correlates with recombinationfrequency. Marker loci are themselves traits and can be assessedaccording to standard linkage analysis by tracking the marker lociduring segregation. Thus, in the context of the present invention, onecM is equal to a 1% chance that a marker locus will be separated fromanother locus (which can be any other trait, e.g., another marker locus,or another trait locus that encodes a QTL), due to crossing over in asingle generation.

When referring to the relationship between two genetic elements, such asa genetic element contributing to resistance and a proximal marker,“coupling” phase linkage indicates the state where the “favorable”allele at the resistance locus is physically associated on the samechromosome strand as the “favorable” allele of the respective linkedmarker locus. In coupling phase, both favorable alleles are inheritedtogether by progeny that inherit that chromosome strand. In “repulsion”phase linkage, the “favorable” allele at the locus of interest (e.g., aQTL for resistance) is physically linked with an “unfavorable” allele atthe proximal marker locus, and the two “favorable” alleles are notinherited together (i.e., the two loci are “out of phase” with eachother).

Genetic Mapping

A “genetic map” is the relationship of genetic linkage among loci on oneor more chromosomes (or linkage groups) within a given species,generally depicted in a diagrammatic or tabular form. “Genetic mapping”is the process of defining the linkage relationships of loci through theuse of genetic markers, populations segregating for the markers, andstandard genetic principles of recombination frequency. A “genetic maplocation” is a location on a genetic map relative to surrounding geneticmarkers on the same linkage group where a specified marker can be foundwithin a given species. In contrast, a physical map of the genome refersto absolute distances (for example, measured in base pairs or isolatedand overlapping contiguous genetic fragments, e.g., contigs). A physicalmap of the genome does not take into account the genetic behavior (e.g.,recombination frequencies) between different points on the physical map.A “genetic recombination frequency” is the frequency of a crossing overevent (recombination) between two genetic loci. Recombination frequencycan be observed by following the segregation of markers and/or traitsfollowing meiosis. In some cases, two different markers can have thesame genetic map coordinates. In that case, the two markers are in suchclose proximity to each other that recombination occurs between themwith such low frequency that it is undetected.

Genetic maps are graphical representations of genomes (or a portion of agenome such as a single chromosome) where the distances between markersare measured by the recombination frequencies between them. Plantbreeders use genetic maps of molecular markers to increase breedingefficiency through MAS, a process where selection for a trait ofinterest is not based on the trait itself but rather on the genotype ofa marker linked to the trait. A molecular marker that demonstratesreliable linkage with a phenotypic trait provides a useful tool forindirectly selecting the trait in a plant population, especially whenaccurate phenotyping is difficult, slow, or expensive.

In general, the closer two markers or genomic loci are on the geneticmap, the closer they lie to one another on the physical map. A lack ofprecise proportionality between cM distances and physical distances canexist due to the fact that the likelihood of genetic recombination isnot uniform throughout the genome; some chromosome regions arecross-over “hot spots,” while other regions demonstrate only rarerecombination events, if any.

Genetic mapping variability can also be observed between differentpopulations of the same crop species. In spite of this variability inthe genetic map that may occur between populations, genetic map andmarker information derived from one population generally remains usefulacross multiple populations in identification of plants with desiredtraits, counter-selection of plants with undesirable traits and inguiding MAS.

As one of skill in the art will recognize, recombination frequencies(and as a result, genetic map positions) in any particular populationare not static. The genetic distances separating two markers (or amarker and a QTL) can vary depending on how the map positions aredetermined. For example, variables such as the parental mappingpopulations used, the software used in the marker mapping or QTLmapping, and the parameters input by the user of the mapping softwarecan contribute to the QTL marker genetic map relationships. However, itis not intended that the invention be limited to any particular mappingpopulations, use of any particular software, or any particular set ofsoftware parameters to determine linkage of a particular marker orchromosome interval with the disease resistance phenotype. It is wellwithin the ability of one of ordinary skill in the art to extrapolatethe novel features described herein to any gene pool or population ofinterest, and using any particular software and software parameters.Indeed, observations regarding genetic markers and chromosome intervalsin populations in addition to those described herein are readily madeusing the teaching of the present disclosure.

Association Mapping

Association or LD mapping techniques aim to identify genotype-phenotypeassociations that are significant. It is effective for fine mapping inoutcrossing species where frequent recombination among heterozygotes canresult in rapid LD decay. LD is non-random association of alleles in acollection of individuals, reflecting the recombinational history ofthat region. Thus, LD decay averages can help determine the number ofmarkers necessary for a genome-wide association study to generate agenetic map with a desired level of resolution.

Large populations are better for detecting recombination, while olderpopulations are generally associated with higher levels of polymorphism,both of which contribute to accelerated LD decay. However, smallereffective population sizes tend to show slower LD decay, which canresult in more extensive haplotype conservation. Understanding of therelationships between polymorphism and recombination is useful indeveloping strategies for efficiently extracting information from theseresources. Association analyses compare the plants' phenotypic scorewith the genotypes at the various loci. Subsequently, any suitable maizegenetic map (for example, a composite map) can be used to help observedistribution of the identified QTL markers and/or QTL marker clusteringusing previously determined map locations of the markers.

Marker Assisted Selection

“Introgression” refers to the transmission of a desired allele of agenetic locus from one genetic background to another. For example,introgression of a desired allele at a specified locus can betransmitted to at least one progeny via a sexual cross between twoparents of the same species, where at least one of the parents has thedesired allele in its genome. Alternatively, for example, transmissionof an allele can occur by recombination between two donor genomes, e.g.,in a fused protoplast, where at least one of the donor protoplasts hasthe desired allele in its genome. The desired allele can be, e.g., aselected allele of a marker, a QTL, a transgene, or the like. In anycase, offspring comprising the desired allele can be repeatedlybackcrossed to a line having a desired genetic background and selectedfor the desired allele, to result in the allele becoming fixed in aselected genetic background.

A primary motivation for development of molecular markers in cropspecies is the potential for increased efficiency in plant breedingthrough MAS. Genetic markers are used to identify plants that contain adesired genotype at one or more loci, and that are expected to transferthe desired genotype, along with a desired phenotype to their progeny.Genetic markers can be used to identify plants containing a desiredgenotype at one locus, or at several unlinked or linked loci (e.g., ahaplotype), and that would be expected to transfer the desired genotype,along with a desired phenotype to their progeny. The present inventionprovides the means to identify plants that are resistant, exhibitimproved resistance or are susceptible to ASR infection by identifyingplants having a specified allele that is linked to ASR-6.01.

In general, MAS uses polymorphic markers that have been identified ashaving a significant likelihood of co-segregation with a resistancetrait. Such markers are presumed to map near a gene or genes that givethe plant its resistance phenotype, and are considered indicators forthe desired trait, and are termed QTL markers. Plants are tested for thepresence or absence of a desired allele in the QTL marker.

Identification of plants or germplasm that include a marker locus ormarker loci linked to a resistance trait or traits provides a basis forperforming MAS. Plants that comprise favorable markers or favorablealleles are selected for, while plants that comprise markers or allelesthat are negatively correlated with resistance can be selected against.Desired markers and/or alleles can be introgressed into plants having adesired (e.g., elite or exotic) genetic background to produce anintrogressed resistant plant or germplasm. In some aspects, it iscontemplated that a plurality of resistance markers are sequentially orsimultaneous selected and/or introgressed. The combinations ofresistance markers that are selected for in a single plant is notlimited, and can include any combination of markers disclosed herein orany marker linked to the markers disclosed herein, or any markerslocated within the QTL intervals defined herein.

In some embodiments, a disease resistant first corn plant or germplasm(the donor) can be crossed with a second corn plant or germplasm (therecipient, e.g., an elite or exotic corn, depending on characteristicsthat are desired in the progeny) to create an introgressed corn plant orgermplasm as part of a breeding program designed to improve diseaseresistance of the recipient corn plant or germplasm. In some aspects,the recipient plant can also contain one or more disease resistant loci,which can be qualitative or quantitative trait loci. In another aspect,the recipient plant can contain a transgene.

In some embodiments, the recipient corn plant or germplasm willtypically display reduced resistance to disease conditions as comparedto the first corn plant or germplasm, while the introgressed corn plantor germplasm will display an increased resistance to disease conditionsas compared to the second plant or germplasm. An introgressed corn plantor germplasm produced by these methods are also a feature of thisinvention.

MAS is a powerful shortcut to selecting for desired phenotypes and forintrogressing desired traits into cultivars (e.g., introgressing desiredtraits into elite lines). MAS is easily adapted to high throughputmolecular analysis methods that can quickly screen large numbers ofplant or germplasm genetic material for the markers of interest and ismuch more cost effective than raising and observing plants for visibletraits.

When a population is segregating for multiple loci affecting one ormultiple traits, e.g., multiple loci involved in resistance, or multipleloci each involved in resistance or tolerance to different diseases, theefficiency of MAS compared to phenotypic screening becomes even greater,because all of the loci can be evaluated in the lab together from asingle sample of DNA.

Introgression of ASR Resistance Loci Using MAS

The introgression of one or more desired loci from a donor line intoanother is achieved via repeated backcrossing to a recurrent parentaccompanied by selection to retain one or more ASR resistance loci fromthe donor parent. Markers associated with ASR resistance are assayed inprogeny and those progeny with one or more ASR resistance markers areselected for advancement. In another aspect, one or more markers can beassayed in the progeny to select for plants with the genotype of theagronomically elite parent. This invention anticipates that traitintrogression activities will require more than one generation, whereinprogeny are crossed to the recurrent (agronomically elite) parent orselfed. Selections are made based on the presence of one or more ASRresistance markers and can also be made based on the recurrent parentgenotype, wherein screening is performed on a genetic marker and/orphenotype basis. In another embodiment, markers of this invention can beused in conjunction with other markers, ideally at least one on eachchromosome of the corn genome, to track the introgression of ASRresistance loci into elite germplasm. In another embodiment, QTLsassociated with ASR resistance will be useful in conjunction with SNPmolecular markers of the present invention to combine quantitative andqualitative ASR resistance in the same plant. It is within the scope ofthis invention to utilize the methods and compositions for traitintegration of ASR resistance. It is contemplated by the inventors thatthe present invention will be useful for developing commercial varietieswith ASR resistance and an agronomically elite phenotype.

In an aspect, this invention could be used on any plant. In anotheraspect, the plant is selected from the genus Zea. In another aspect, theplant is selected from the species Zea mays. In a further aspect, theplant is selected from the subspecies Zea mays L. ssp. mays. In anadditional aspect, the plant is selected from the group Zea mays L.subsp. mays Indentata, otherwise known as dent corn. In another aspect,the plant is selected from the group Zea mays L. subsp. mays Indurata,otherwise known as flint corn. In an aspect, the plant is selected fromthe group Zea mays L. subsp. mays Saccharata, otherwise known as sweetcorn. In another aspect, the plant is selected from the group Zea maysL. subsp. mays Amylacea, otherwise known as flour corn. In a furtheraspect, the plant is selected from the group Zea mays L. subsp. maysEverta, otherwise known as pop corn. Zea plants include hybrids,inbreds, partial inbreds, or members of defined or undefinedpopulations.

In another aspect, a corn plant of the invention can show a comparativeresistance compared to a non-resistant control corn plant. In thisaspect, a control corn plant will preferably be genetically similarexcept for the disease resistance allele or alleles in question. Suchplants can be grown under similar conditions with equivalent or nearequivalent exposure to the pathogen.

Transgenic Plants Transformation Constructs

Vectors used for plant transformation may include, for example,plasmids, cosmids, YACs (yeast artificial chromosomes), BACs (bacterialartificial chromosomes) or any other suitable cloning system, as well asfragments of DNA therefrom. Thus when the term “vector” or “expressionvector” is used, all of the foregoing types of vectors, as well asnucleic acid sequences isolated therefrom, are included. It iscontemplated that utilization of cloning systems with large insertcapacities will allow introduction of large DNA sequences comprisingmore than one selected gene. In accordance with the present disclosure,this could be used to introduce genes corresponding to, e.g., an entirebiosynthetic pathway, into a plant.

Particularly useful for transformation are expression cassettes whichhave been isolated from such vectors. DNA segments used for transformingplant cells will generally comprise the cDNA, gene, or genes which onedesires to introduce into and have expressed in the host cells. TheseDNA segments can further include structures such as promoters,enhancers, polylinkers, or regulatory genes as desired. The DNA segmentor gene chosen for cellular introduction will often encode a proteinwhich will be expressed in the resultant recombinant cells resulting ina screenable or selectable trait and/or which will impart an improvedphenotype to the resulting transgenic plant.

Regulatory elements such as promoters, leaders, enhancers, introns, andtranscription termination regions (or 3′ UTRs) can play an integral partin the overall expression of genes in living cells. The term “regulatoryelement,” as used herein, refers to a DNA molecule havinggene-regulatory activity. The term “gene-regulatory activity,” as usedherein, refers to the ability to affect the expression of an operablylinked transcribable DNA molecule, for instance by affecting thetranscription and/or translation of the operably linked transcribableDNA molecule. Regulatory elements, such as promoters, leaders,enhancers, and introns that function in plants are therefore useful formodifying plant phenotypes through genetic engineering.

As used herein, the term “intron” refers to a DNA molecule that may beisolated or identified from the genomic copy of a gene and may bedefined generally as a region spliced out during messenger RNA (mRNA)processing prior to translation. Alternately, an intron may be asynthetically produced or manipulated DNA element. An intron may containenhancer elements that effect the transcription of operably linkedgenes. An intron may be used as a regulatory element for modulatingexpression of an operably linked transcribable DNA molecule. A constructmay comprise an intron, and the intron may or may not be heterologouswith respect to the transcribable DNA molecule. Examples of introns inthe art include the rice actin intron and the corn HSP70 intron. Inplants, the inclusion of some introns in constructs leads to increasedmRNA and protein accumulation relative to constructs lacking the intron.This effect has been termed “intron mediated enhancement” (IME) of geneexpression. Introns known to stimulate expression in plants have beenidentified in maize genes (e.g., tubA1, Adh1, Sh1, and Ubi1), in ricegenes (e.g., tpi) and in dicotyledonous plant genes like those fromPetunia (e.g., rbcS), potato (e.g., st-ls1) and from Arabidopsisthaliana (e.g., ubq3 and pat1). It has been shown that deletions ormutations within the splice sites of an intron reduce gene expression,indicating that splicing might be needed for IME. However, that splicingper se is not required, as IME in dicotyledonous plants has been shownby point mutations within the splice sites of the pat1 gene from A.thaliana. Multiple uses of the same intron in one plant have been shownto exhibit disadvantages. In those cases, it is necessary to have acollection of basic control elements for the construction of appropriaterecombinant DNA elements.

As used herein, the term “enhancer” or “enhancer element” refers to acis-acting regulatory element, a.k.a. cis-element, which confers anaspect of the overall expression pattern, but is usually insufficientalone to drive transcription, of an operably linked DNA sequence. Unlikepromoters, enhancer elements do not usually include a transcriptionstart site (TSS) or TATA box or equivalent DNA sequence. A promoter orpromoter fragment may naturally comprise one or more enhancer elementsthat affect the transcription of an operably linked DNA sequence. Anenhancer element may also be fused to a promoter to produce a chimericpromoter cis-element, which confers an aspect of the overall modulationof gene expression.

Regulatory elements may be characterized by their gene expressionpattern, e.g., positive and/or negative effects, such as constitutiveexpression or temporal, spatial, developmental, tissue, environmental,physiological, pathological, cell cycle, and/or chemically responsiveexpression, and any combination thereof, as well as by quantitative orqualitative indications. As used herein, a “gene expression pattern” isany pattern of transcription of an operably linked DNA molecule into atranscribed RNA molecule. The transcribed RNA molecule may be translatedto produce a protein molecule or may provide an antisense or otherregulatory RNA molecule, such as a double-stranded RNA (dsRNA), atransfer RNA (tRNA), a ribosomal RNA (rRNA), a microRNA (miRNA), and thelike.

As used herein, the term “protein expression” is any pattern oftranslation of a transcribed RNA molecule into a protein molecule.Protein expression may be characterized by its temporal, spatial,developmental, or morphological qualities, as well as by quantitative orqualitative indications.

A promoter is useful as a regulatory element for modulating theexpression of an operably linked transcribable DNA molecule. As usedherein, the term “promoter” refers generally to a DNA molecule that isinvolved in recognition and binding of RNA polymerase II and otherproteins, such as trans-acting transcription factors, to initiatetranscription. A promoter may originate from the 5′ untranslated region(5′ UTR) of a gene. Alternately, promoters may be synthetically producedor manipulated DNA molecules. Promoters may also be chimeric. As usedherein, the term “chimeric” refers to a single DNA molecule produced byfusing a first DNA molecule to a second DNA molecule, where neither thefirst nor the second DNA molecule would normally be contained in thatconfiguration, i.e., fused to the other. The chimeric DNA molecule isthus a new DNA molecule not otherwise normally contained in nature. Asused herein, the term “chimeric promoter” refers to a promoter producedthrough such manipulation of DNA molecules. A chimeric promoter maycombine two or more DNA fragments, for example, the fusion of a promoterto an enhancer element. Thus, the design, construction, and use ofchimeric promoters according to the methods disclosed herein formodulating the expression of operably linked transcribable DNA moleculesare encompassed by the disclosure.

In specific embodiments, chimeric DNA molecules and any variants orderivatives thereof as described herein, are further defined ascomprising promoter activity, i.e., are capable of acting as a promoterin a host cell, such as in a transgenic plant. In still further specificembodiments, a fragment may be defined as exhibiting promoter activitypossessed by the starting promoter molecule from which it is derived, ora fragment may comprise a “minimal promoter” which provides a basallevel of transcription and is comprised of a TATA box or equivalent DNAsequence for recognition and binding of the RNA polymerase II complexfor initiation of transcription.

Exemplary promoters for expression of a nucleic acid sequence includeplant promoters such as the CaMV 35S promoter, or others such as CaMV19S, nos, Adh, sucrose synthase, α-tubulin, actin, cab, PEPCase or thosepromoters associated with the R gene complex. Tissue-specific promoterssuch as leaf specific promoters, or tissue selective promoters (e.g.,promoters that direct greater expression in leaf primordia than in othertissues), and tissue-specific enhancers are also contemplated to beuseful, as are inducible promoters such as ABA- and turgor-induciblepromoters. Any suitable promoters known in the art may be used toexpress defensin or defensin-like coding sequences in a plant. In anembodiment, the CaMV35S promoter may be used to express defensin ordefensin-like coding sequences in a plant. In yet another embodiment, adisease or pathogen inducible promoter can be used to express defensinor defensin like proteins. Examples of disease or pathogen induciblepromoters can be found in Kooshki et al. Plant Science 165 (2003)213-219, Koschmann et al. Plant Physiology 160 (2012) 178-191, Rushtonet al. The Plant Cell, 14 (2002) 749-762, and Kirsch et al. The PlantJournal (2001) 26 217-227.

The DNA sequence between the transcription initiation site and the startof the coding sequence, i.e., the untranslated leader sequence, can alsoinfluence gene expression. As used herein, the term “leader” refers to aDNA molecule from the untranslated 5′ region (5′ UTR) of a gene anddefined generally as a DNA segment between the transcription start site(TSS) and the protein coding sequence start site. Alternately, leadersmay be synthetically produced or manipulated DNA elements. A leader canbe used as a 5′ regulatory element for modulating expression of anoperably linked transcribable DNA molecule. Leader molecules may be usedwith a heterologous promoter or with their native promoter. One may thuswish to employ a particular leader sequence with a transformationconstruct of the present disclosure. In an embodiment, leader sequencesare contemplated to include those which comprise sequences predicted todirect optimum expression of the attached gene, i.e., to include aconsensus leader sequence which may increase or maintain mRNA stabilityand prevent inappropriate initiation of translation. The choice of suchsequences will be known to those of skill in the art in light of thepresent disclosure. In some embodiments, sequences that are derived fromgenes that are highly expressed in plants may be used for expression ofdefensin or defensin-like coding sequences.

It is envisioned that defensin or defensin-like coding sequences may beintroduced under the control of novel promoters, enhancers, etc., orhomologous or tissue-specific or tissue-selective, or pathogen ordisease promoters or control elements. Vectors for use intissue-specific targeting of genes in transgenic plants will typicallyinclude tissue-specific or tissue-selective promoters and may alsoinclude other tissue-specific or tissue-selective control elements suchas enhancer sequences. Promoters which direct specific or enhancedexpression in certain plant tissues will be known to those of skill inthe art in light of the present disclosure.

Transformation constructs prepared in accordance with the presentdisclosure may further include a 3′ end DNA sequence that acts as asignal to terminate transcription and allow for the polyadenylation ofthe mRNA produced by coding sequences operably linked to a promoter. Asused herein, the term “3′ transcription termination molecule,” “3′untranslated region” or “3′ UTR” herein refers to a DNA molecule that isused during transcription to the untranslated region of the 3′ portionof an mRNA molecule. The 3′ untranslated region of an mRNA molecule maybe generated by specific cleavage and 3′ polyadenylation, also known asa polyA tail. A 3′ UTR may be operably linked to and located downstreamof a transcribable DNA molecule and may include a polyadenylation signaland other regulatory signals capable of affecting transcription, mRNAprocessing, or gene expression. PolyA tails are thought to function inmRNA stability and in initiation of translation. Examples of 3′transcription termination molecules in the art are the nopaline synthase3′ region; wheat hsp17 3′ region, pea rubisco small subunit 3′ region,cotton E6 3′ region, and the coixin 3′ UTR.

3′ UTRs typically find beneficial use for the recombinant expression ofspecific DNA molecules. A weak 3′ UTR has the potential to generateread-through, which may affect the expression of the DNA moleculelocated in the neighboring expression cassettes. Appropriate control oftranscription termination can prevent read-through into DNA sequences(e.g., other expression cassettes) localized downstream and can furtherallow efficient recycling of RNA polymerase to improve gene expression.Efficient termination of transcription (release of RNA Polymerase IIfrom the DNA) is prerequisite for re-initiation of transcription andthereby directly affects the overall transcript level. Subsequent totranscription termination, the mature mRNA is released from the site ofsynthesis and template transported to the cytoplasm. Eukaryotic mRNAsare accumulated as poly(A) forms in vivo, making it difficult to detecttranscriptional termination sites by conventional methods. However,prediction of functional and efficient 3′ UTRs by bioinformatics methodsis difficult in that there are no conserved DNA sequences that wouldallow easy prediction of an effective 3′ UTR. In one embodiment, thenative terminator of a defensin or defensin-like coding sequence may beused. Alternatively, a heterologous 3′ end may enhance the expression ofsense or antisense defensin or defensin-like coding sequences.

Sequences that are joined to the coding sequence of an expressed gene,which are removed post-translationally from the initial translationproduct and which facilitate the transport of the protein into orthrough intracellular or extracellular membranes, are termed transit ortargeting peptide (usually into vacuoles, vesicles, plastids and otherintracellular organelles) and signal peptide or sequences (usually tothe endoplasmic reticulum, Golgi apparatus, and outside of the cellularmembrane). By facilitating the transport of the protein intocompartments inside and outside the cell, these sequences may increasethe accumulation of gene products by protecting them from proteolyticdegradation. These sequences also allow for additional mRNA sequencesfrom highly expressed genes to be attached to the coding sequence of thegenes. Since mRNA being translated by ribosomes is more stable thannaked mRNA, the presence of translatable mRNA in front of the gene mayincrease the overall stability of the mRNA transcript from the gene andthereby increase synthesis of the gene product. Since transit and signalsequences are usually post-translationally removed from the initialtranslation product, the use of these sequences allows for the additionof extra translated sequences that may not appear on the finalpolypeptide. It further is contemplated that targeting of certainproteins may be desirable in order to enhance the stability of theprotein.

Additionally, vectors may be constructed and employed in theintracellular targeting of a specific gene product within the cells of atransgenic plant or in directing a protein to the extracellularenvironment. This generally will be achieved by joining a DNA sequenceencoding a transit or signal peptide sequence to the coding sequence ofa particular gene. The resultant transit or signal peptide willtransport the protein to a particular intracellular or extracellulardestination, respectively, and will then be post-translationallyremoved.

By employing a selectable or screenable marker, one can provide orenhance the ability to identify transformants. “Marker genes” are genesthat impart a distinct phenotype to cells expressing the marker proteinand thus allow such transformed cells to be distinguished from cellsthat do not have the marker. Such genes may encode either a selectableor screenable marker, depending on whether the marker confers a traitwhich one can “select” for by chemical means, i.e., through the use of aselective agent (e.g., a herbicide, antibiotic, or the like), or whetherit is simply a trait that one can identify through observation ortesting, i.e., by “screening” (e.g., the green fluorescent protein). Ofcourse, many examples of suitable marker proteins are known to the artand can be employed in the practice of the present disclosure.

Selectable marker transgenes may also be used with the presentdisclosure. As used herein the term “selectable marker transgene” refersto any transcribable DNA molecule whose expression in a transgenicplant, tissue or cell, or lack thereof, can be screened for or scored insome way. Selectable marker genes, and their associated selection andscreening techniques, for use in the practice of the present disclosureare known in the art and include, but are not limited to, transcribableDNA molecules encoding β-glucuronidase (GUS), green fluorescent protein(GFP), proteins that confer antibiotic resistance, and proteins thatconfer herbicide resistance

Plant Cell Transformation Methods

Numerous methods for transforming chromosomes in a plant cell withrecombinant DNA are known in the art and are used in methods ofproducing a transgenic plant cell and plant. Two effective methods forsuch transformation are Agrobacterium-mediated transformation andmicroprojectile bombardment-mediated transformation. Microprojectilebombardment methods are illustrated in U.S. Pat. No. 5,015,580(soybean); U.S. Pat. No. 5,550,318 (corn); U.S. Pat. No. 5,538,880(corn); U.S. Pat. No. 5,914,451 (soybean); U.S. Pat. No. 6,160,208(corn); U.S. Pat. No. 6,399,861 (corn); U.S. Pat. No. 6,153,812 (wheat)and U.S. Pat. No. 6,365,807 (rice). Agrobacterium-mediatedtransformation methods are described in U.S. Pat. No. 5,159,135(cotton); U.S. Pat. No. 5,824,877 (soybean); U.S. Pat. No. 5,463,174(canola); U.S. Pat. No. 5,591,616 (corn); U.S. Pat. No. 5,846,797(cotton); U.S. Pat. No. 6,384,301 (soybean), U.S. Pat. No. 7,026,528(wheat) and U.S. Pat. No. 6,329,571 (rice), and US Patent ApplicationPublication Nos. US 2004/0087030 A1 (cotton), and US 2001/0042257 A1(sugar beet), all of which are incorporated herein by reference in theirentirety. Transformation of plant material is practiced in tissueculture on nutrient media, for example a mixture of nutrients that allowcells to grow in vitro. Recipient cell targets include, but are notlimited to, meristem cells, shoot tips, hypocotyls, calli, immature ormature embryos, and gametic cells such as microspores, pollen, sperm andegg cells. Callus can be initiated from tissue sources including, butnot limited to, immature or mature embryos, hypocotyls, seedling apicalmeristems, microspores and the like. Cells containing a transgenicnucleus are grown into transgenic plants.

In addition to direct transformation of a plant material with arecombinant DNA, a transgenic plant can be prepared by crossing a firstplant comprising a recombinant DNA with a second plant lacking therecombinant DNA. For example, recombinant DNA can be introduced into afirst plant line that is amenable to transformation, which can becrossed with a second plant line to introgress the recombinant DNA intothe second plant line. A transgenic plant with recombinant DNA providingan enhanced trait, for example, enhanced yield, can be crossed with atransgenic plant line having another recombinant DNA that confersanother trait, for example herbicide resistance or pest resistance orenhanced water use efficiency, to produce progeny plants havingrecombinant DNA that confers both traits. Typically, in such breedingfor combining traits the transgenic plant donating the additional traitis the male line and the transgenic plant carrying the base traits isthe female line. The progeny of this cross will segregate such that someof the plants will carry the DNA for both parental traits and some willcarry DNA for one parental trait; such plants can be identified bymarkers associated with parental recombinant DNA, for example, markeridentification by analysis for recombinant DNA or, in the case where aselectable marker is linked to the recombinant DNA, by application usinga selective agent such as a herbicide for use with a herbicideresistance marker, or by selection for the enhanced trait. Progenyplants carrying DNA for both parental traits can be crossed back intothe female parent line multiple times, for example usually 6 to 8generations, to produce a progeny plant with substantially the samegenotype as the original transgenic parental line but for therecombinant DNA of the other transgenic parental line.

In transformation, DNA is typically introduced into only a smallpercentage of target plant cells in any one transformation experiment.Marker genes are used to provide an efficient system for identificationof those cells that are stably transformed by receiving and integratinga recombinant DNA molecule into their genomes. Preferred marker genesprovide selective markers which confer resistance to a selective agent,such as an antibiotic or an herbicide. Any of the herbicides to whichplants of this disclosure can be resistant is an agent for selectivemarkers. Potentially transformed cells are exposed to the selectiveagent. In the population of surviving cells are those cells where,generally, the resistance-conferring gene is integrated and expressed atsufficient levels to permit cell survival. Cells can be tested furtherto confirm stable integration of the exogenous DNA. Commonly usedselective marker genes include those conferring resistance toantibiotics such as kanamycin and paromomycin (nptII), hygromycin B (aphIV), spectinomycin (aadA) and gentamycin (aac3 and aacC4) or resistanceto herbicides such as glufosinate (bar or pat), dicamba (DMO) andglyphosate (aroA or EPSPS). Examples of such selectable markers areillustrated in U.S. Pat. Nos. 5,550,318; 5,633,435; 5,780,708 and6,118,047. Markers which provide an ability to visually screentransformants can also be employed, for example, a gene expressing acolored or fluorescent protein such as a luciferase or green fluorescentprotein (GFP) or a gene expressing a beta-glucuronidase or uidA gene(GUS) for which various chromogenic substrates are known.

Transgenic Plants and Seeds

Transgenic plants derived from transgenic plant cells having atransgenic nucleus of this disclosure are grown to generate transgenicplants having an enhanced trait as compared to a control plant, andproduce transgenic seed and haploid pollen of this disclosure. Suchplants with enhanced traits are identified by selection of transformedplants or progeny seed for the enhanced trait. For efficiency aselection method is designed to evaluate multiple transgenic plants(events) comprising the recombinant DNA, for example multiple plantsfrom 2 to 20 or more transgenic events. Transgenic plants grown fromtransgenic seeds provided herein demonstrate improved agronomic traits,such as resistance to anthracnose stalk rot in maize.

Definitions

The definitions and methods provided define the present invention andguide those of ordinary skill in the art in the practice of the presentinvention. Unless otherwise noted, terms are to be understood accordingto conventional usage by those of ordinary skill in the relevant art.Examples of resources describing many of the terms related to molecularbiology used herein can be found in in Alberts et al., Molecular Biologyof The Cell, 5^(th) Edition, Garland Science Publishing, Inc.: New York,2007; Rieger et al., Glossary of Genetics: Classical and Molecular, 5thedition, Springer-Verlag: New York, 1991; King et al, A Dictionary ofGenetics, 6th ed, Oxford University Press: New York, 2002; and Lewin,Genes Icorn, Oxford University Press: New York, 2007. The nomenclaturefor DNA bases as set forth at 37 CFR § 1.822 is used.

“Adjacent”, when used to describe a nucleic acid molecule thathybridizes to DNA containing a polymorphism, refers to a nucleic acidthat hybridizes to DNA sequences that directly abut the polymorphicnucleotide base position. For example, a nucleic acid molecule that canbe used in a single base extension assay is “adjacent” to thepolymorphism.

“Allele” refers to an alternative nucleic acid sequence at a particularlocus; the length of an allele can be as small as 1 nucleotide base, butis typically larger. For example, a first allele can occur on onechromosome, while a second allele occurs on a second homologouschromosome, e.g., as occurs for different chromosomes of a heterozygousindividual, or between different homozygous or heterozygous individualsin a population. A favorable allele is the allele at a particular locusthat confers, or contributes to, an agronomically desirable phenotype,or alternatively, is an allele that allows the identification ofsusceptible plants that can be removed from a breeding program orplanting. A favorable allele of a marker is a marker allele thatsegregates with the favorable phenotype, or alternatively, segregateswith susceptible plant phenotype, therefore providing the benefit ofidentifying disease prone plants. A favorable allelic form of achromosome interval is a chromosome interval that includes a nucleotidesequence that contributes to superior agronomic performance at one ormore genetic loci physically located on the chromosome interval. “Allelefrequency” refers to the frequency (proportion or percentage) at whichan allele is present at a locus within an individual, within a line, orwithin a population of lines. For example, for an allele “A,” diploidindividuals of genotype “AA,” “Aa,” or “aa” have allele frequencies of1.0, 0.5, or 0.0, respectively. One can estimate the allele frequencywithin a line by averaging the allele frequencies of a sample ofindividuals from that line. Similarly, one can calculate the allelefrequency within a population of lines by averaging the allelefrequencies of lines that make up the population. For a population witha finite number of individuals or lines, an allele frequency can beexpressed as a count of individuals or lines (or any other specifiedgrouping) containing the allele. An allele positively correlates with atrait when it is linked to it and when presence of the allele is anindictor that the desired trait or trait form will occur in a plantcomprising the allele. An allele negatively correlates with a trait whenit is linked to it and when presence of the allele is an indicator thata desired trait or trait form will not occur in a plant comprising theallele.

“Crossed” or “cross” means to produce progeny via fertilization (e.g.cells, seeds or plants) and includes crosses between plants (sexual) andself fertilization (selfing).

“Elite line” means any line that has resulted from breeding andselection for superior agronomic performance. Numerous elite lines areavailable and known to those of skill in the art of corn breeding. An“elite population” is an assortment of elite individuals or lines thatcan be used to represent the state of the art in terms of agronomicallysuperior genotypes of a given crop species, such as corn. Similarly, an“elite germplasm” or elite strain of germplasm is an agronomicallysuperior germplasm.

“Exogenous nucleic acid” is a nucleic acid that is not native to aspecified system (e.g., a germplasm, plant, variety, etc.), with respectto sequence, genomic position, or both. As used herein, the terms“exogenous” or “heterologous” as applied to polynucleotides orpolypeptides typically refers to molecules that have been artificiallysupplied to a biological system (e.g., a plant cell, a plant gene, aparticular plant species or variety or a plant chromosome under study)and are not native to that particular biological system. The terms canindicate that the relevant material originated from a source other thana naturally occurring source, or can refer to molecules having anon-natural configuration, genetic location or arrangement of parts. Incontrast, for example, a “native” or “endogenous” gene is a gene thatdoes not contain nucleic acid elements encoded by sources other than thechromosome or other genetic element on which it is normally found innature. An endogenous gene, transcript or polypeptide is encoded by itsnatural chromosomal locus, and not artificially supplied to the cell.

“Genetic element” or “gene” refers to a heritable sequence of DNA, i.e.,a genomic sequence, with functional significance. The term “gene” canalso be used to refer to, e.g., a cDNA and/or a mRNA encoded by agenomic sequence, as well as to that genomic sequence.

“Genotype” is the genetic constitution of an individual (or group ofindividuals) at one or more genetic loci, as contrasted with theobservable trait (the phenotype). Genotype is defined by the allele(s)of one or more known loci that the individual has inherited from itsparents. The term genotype can be used to refer to an individual'sgenetic constitution at a single locus, at multiple loci, or, moregenerally, the term genotype can be used to refer to an individual'sgenetic make-up for all the genes in its genome. A “haplotype” is thegenotype of an individual at a plurality of genetic loci. Typically, thegenetic loci described by a haplotype are physically and geneticallylinked, i.e., on the same chromosome interval. The terms “phenotype,” or“phenotypic trait” or “trait” refers to one or more trait of anorganism. The phenotype can be observable to the naked eye, or by anyother means of evaluation known in the art, e.g., microscopy,biochemical analysis, genomic analysis, an assay for a particulardisease resistance, etc. In some cases, a phenotype is directlycontrolled by a single gene or genetic locus, i.e., a “single genetrait.” In other cases, a phenotype is the result of several genes.

“Germplasm” refers to genetic material of or from an individual (e.g., aplant), a group of individuals (e.g., a plant line, variety or family),or a clone derived from a line, variety, species, or culture. Thegermplasm can be part of an organism or cell, or can be separate fromthe organism or cell. In general, germplasm provides genetic materialwith a specific molecular makeup that provides a physical foundation forsome or all of the hereditary qualities of an organism or cell culture.As used herein, germplasm includes cells, seed or tissues from which newplants may be grown, or plant parts, such as leaves, stems, pollen, orcells that can be cultured into a whole plant.

“Linkage disequilibrium” refers to a non-random segregation of geneticloci or traits (or both). In either case, linkage disequilibrium impliesthat the relevant loci are within sufficient physical proximity along alength of a chromosome so that they segregate together with greater thanrandom (i.e., non-random) frequency (in the case of co-segregatingtraits, the loci that underlie the traits are in sufficient proximity toeach other). Linked loci co-segregate more than 50% of the time, e.g.,from about 51% to about 100% of the time. The term “physically linked”is sometimes used to indicate that two loci, e.g., two marker loci, arephysically present on the same chromosome. Advantageously, the twolinked loci are located in close proximity such that recombinationbetween homologous chromosome pairs does not occur between the two lociduring meiosis with high frequency, e.g., such that linked locicosegregate at least about 90% of the time, e.g., 91%, 92%, 93%, 94%,95%, 96%, 97%, 98%, 99%, 99.5%, 99.75%, or more of the time.

“Locus” a chromosome region where a polymorphic nucleic acid, traitdeterminant, gene or marker is located. The loci of this inventioncomprise one or more polymorphisms in a population; i.e., alternativealleles are present in some individuals. A “gene locus” is a specificchromosome location in the genome of a species where a specific gene canbe found.

“Marker Assay” means a method for detecting a polymorphism at aparticular locus using a particular method, e.g. measurement of at leastone phenotype (such as seed color, flower color, or other visuallydetectable trait), restriction fragment length polymorphism (RFLP),single base extension, electrophoresis, sequence alignment, allelicspecific oligonucleotide hybridization (ASO), random amplifiedpolymorphic DNA (RAPD), microarray-based technologies, and nucleic acidsequencing technologies, etc. “Marker Assisted Selection” (MAS) is aprocess by which phenotypes are selected based on marker genotypes.

“Molecular phenotype” is a phenotype detectable at the level of apopulation of one or more molecules. Such molecules can be nucleicacids, proteins, or metabolites. A molecular phenotype could be anexpression profile for one or more gene products, e.g., at a specificstage of plant development, in response to an environmental condition orstress, etc.

“Operably linked” refers to the association of two or more nucleic acidelements in a recombinant DNA construct, e.g. as when a promoter isoperably linked with DNA that is transcribed to RNA whether forexpressing or suppressing a protein. Recombinant DNA constructs can bedesigned to express a protein which can be an endogenous protein, anexogenous homologue of an endogenous protein or an exogenous proteinwith no native homologue. Alternatively, recombinant DNA constructs canbe designed to suppress the level of an endogenous protein, e.g. bysuppression of the native gene. Such gene suppression can be effectivelyemployed through a native RNA interference (RNAi) mechanism in whichrecombinant DNA comprises both sense and anti-sense oriented DNA matchedto the gene targeted for suppression where the recombinant DNA istranscribed into RNA that can form a double-strand to initiate an RNAimechanism. Gene suppression can also be effected by recombinant DNA thatcomprises anti-sense oriented DNA matched to the gene targeted forsuppression. Gene suppression can also be effected by recombinant DNAthat comprises DNA that is transcribed to a microRNA matched to the genetargeted for suppression.

“Percent identity” or “% identity” means the extent to which twooptimally aligned DNA or protein segments are invariant throughout awindow of alignment of components, for example nucleotide sequence oramino acid sequence. An “identity fraction” for aligned segments of atest sequence and a reference sequence is the number of identicalcomponents that are shared by sequences of the two aligned segmentsdivided by the total number of sequence components in the referencesegment over a window of alignment which is the smaller of the full testsequence or the full reference sequence.

“Phenotype” means the detectable characteristics of a cell or organismwhich can be influenced by genotype.

“Plant” refers to a whole plant any part thereof, or a cell or tissueculture derived from a plant, comprising any of: whole plants, plantcomponents or organs (e.g., leaves, stems, roots, etc.), plant tissues,seeds, plant cells, and/or progeny of the same. A plant cell is abiological cell of a plant, taken from a plant or derived throughculture from a cell taken from a plant.

“Polymorphism” means the presence of one or more variations in apopulation. A polymorphism may manifest as a variation in the nucleotidesequence of a nucleic acid or as a variation in the amino acid sequenceof a protein. Polymorphisms include the presence of one or morevariations of a nucleic acid sequence or nucleic acid feature at one ormore loci in a population of one or more individuals. The variation maycomprise but is not limited to one or more nucleotide base changes, theinsertion of one or more nucleotides or the deletion of one or morenucleotides. A polymorphism may arise from random processes in nucleicacid replication, through mutagenesis, as a result of mobile genomicelements, from copy number variation and during the process of meiosis,such as unequal crossing over, genome duplication and chromosome breaksand fusions. The variation can be commonly found or may exist at lowfrequency within a population, the former having greater utility ingeneral plant breeding and the latter may be associated with rare butimportant phenotypic variation. Useful polymorphisms may include singlenucleotide polymorphisms (SNPs), insertions or deletions in DNA sequence(Indels), simple sequence repeats of DNA sequence (SSRs), a restrictionfragment length polymorphism, and a tag SNP. A genetic marker, a gene, aDNA-derived sequence, a RNA-derived sequence, a promoter, a 5′untranslated region of a gene, a 3′ untranslated region of a gene,microRNA, siRNA, a resistance locus, a satellite marker, a transgene,mRNA, ds mRNA, a transcriptional profile, and a methylation pattern mayalso comprise polymorphisms. In addition, the presence, absence, orvariation in copy number of the preceding may comprise polymorphisms.

A “population of plants” or “plant population” means a set comprisingany number, including one, of individuals, objects, or data from whichsamples are taken for evaluation, e.g. estimating QTL effects. Mostcommonly, the terms relate to a breeding population of plants from whichmembers are selected and crossed to produce progeny in a breedingprogram. A population of plants can include the progeny of a singlebreeding cross or a plurality of breeding crosses, and can be eitheractual plants or plant derived material, or in silico representations ofthe plants. The population members need not be identical to thepopulation members selected for use in subsequent cycles of analyses orthose ultimately selected to obtain final progeny plants. Often, a plantpopulation is derived from a single biparental cross, but may alsoderive from two or more crosses between the same or different parents.Although a population of plants may comprise any number of individuals,those of skill in the art will recognize that plant breeders commonlyuse population sizes ranging from one or two hundred individuals toseveral thousand, and that the highest performing 5-20% of a populationis what is commonly selected to be used in subsequent crosses in orderto improve the performance of subsequent generations of the population.

“Resistance” or “improved resistance” in a plant to disease conditionsis an indication that the plant is more able to reduce disease burdenthan a non-resistant or less resistant plant. Resistance is a relativeterm, indicating that a “resistant” plant is more able to reduce diseaseburden compared to a different (less resistant) plant (e.g., a differentcorn line) grown in similar disease conditions. One of skill willappreciate that plant resistance to disease conditions varies widely,and can represent a spectrum of more-resistant or less-resistantphenotypes. However, by simple observation, one of skill can generallydetermine the relative resistance of different plants, plant lines, orplant families under disease conditions, and furthermore, will alsorecognize the phenotypic gradations of “resistant.”

“Resistance locus” means a locus that contributes resistance, tolerance,or susceptibility to anthracnose stalk rot.

“Resistance allele” means the nucleic acid sequence associated withresistance or tolerance to disease.

“Tolerance locus” means a locus associated with tolerance or resistanceto disease. For instance, a tolerance locus according to the presentinvention may, in one embodiment, control tolerance or susceptibilityfor one or more races of Colletotrichum graminicola.

“Tolerance allele” means the nucleic acid sequence associated withtolerance or resistance to disease.

“Recombinant” in reference to a nucleic acid or polypeptide indicatesthat the material (e.g., a recombinant nucleic acid, gene,polynucleotide, polypeptide, etc.) has been altered by humanintervention. The term recombinant can also refer to an organism thatharbors recombinant material, e.g., a plant that comprises a recombinantnucleic acid is considered a recombinant plant.

“Tolerance” or “improved tolerance” in a plant to disease conditions isan indication that the plant is less affected by disease conditions withrespect to yield, survivability and/or other relevant agronomicmeasures, compared to a less resistant, more “susceptible” plant.Tolerance is a relative term, indicating that a “tolerant” plantsurvives and/or produces better yields in disease conditions compared toa different (less tolerant) plant (e.g., a different corn line strain)grown in similar disease conditions. One of skill will appreciate thatplant tolerance to disease conditions varies widely, and can represent aspectrum of more-tolerant or less-tolerant phenotypes. However, bysimple observation, one of skill can generally determine the relativetolerance or susceptibility of different plants, plant lines or plantfamilies under disease conditions, and furthermore, will also recognizethe phenotypic gradations of “tolerant.”

“Transgenic plant” refers to a plant that comprises within its cells aheterologous polynucleotide. Generally, the heterologous polynucleotideis stably integrated within the genome such that the polynucleotide ispassed on to successive generations. The heterologous polynucleotide maybe integrated into the genome alone or as part of a recombinantexpression cassette. “Transgenic” is used herein to refer to any cell,cell line, callus, tissue, plant part or plant, the genotype of whichhas been altered by the presence of heterologous nucleic acid includingthose transgenic organisms or cells initially so altered, as well asthose created by crosses or asexual propagation from the initialtransgenic organism or cell. The term “transgenic” as used herein doesnot encompass the alteration of the genome (chromosomal orextrachromosomal) by conventional plant breeding methods (e.g., crosses)or by naturally occurring events such as random cross-fertilization,non-recombinant viral infection, non-recombinant bacterialtransformation, non-recombinant transposition, or spontaneous mutation.

“Vector” is a polynucleotide or other molecule that transfers nucleicacids between cells. Vectors are often derived from plasmids,bacteriophages, or viruses and optionally comprise parts which mediatevector maintenance and enable its intended use. A “cloning vector” or“shuttle vector” or “subcloning vector” contains operably linked partsthat facilitate subcloning steps (e.g., a multiple cloning sitecontaining multiple restriction endonuclease sites). The term“expression vector” as used herein refers to a vector comprisingoperably linked polynucleotide sequences that facilitate expression of acoding sequence in a particular host organism (e.g., a bacterialexpression vector or a plant expression vector).

“Yield” is the culmination of all agronomic traits as determined by theproductivity per unit area of a particular plant product of commercialvalue. “Agronomic traits,” include the underlying genetic elements of agiven plant variety that contribute to yield over the course of growingseason.

EXAMPLES Example 1. Field Studies A, B and C Biparental MappingPopulations

Parental lines were selected from resistant inbred lines: CV820914,CV094802 and CV594360, and susceptible inbred lines: CV391950, 1294213and 1283669. CV391950 is described in U.S. Pat. Nos. 7,718,859;1,294,213 is described in U.S. Pat. Nos. 7,166,779; and 1,283,669 isdescribed in U.S. Pat. No. 7,414,181. Number of lines derived were 168doubled-haploid from CV820914/CV391950, 180 BC1F3 inbred fromCV094802/1294213*2 and 178 BC1F2 inbred from I283669*2/CV594360. 168hybrid lines were also derived from the cross of (CV820914/CV391950)BC1F3 and testers that are modest-resistant or neutral to ASR (Table 1).

TABLE 1 Mapping populations Field Resistant Susceptible PopulationNumber Study Mapping Population Line Line Type Gender of Lines ACV820914/CV391950 CV820914 CV391950 DH M 168 A CV820914/CV391950CV820914 CV391950 (BC1F3XTester)F1 M 168 B CV094802/I294213*2 CV094802I294213 BC1F3 F 180 C I283669*2/CV594360 CV594360 I283669 BC1F2 M 178

Inoculation and Rating Scale of Phenotypes

Corn plants grown in a field were inoculated 14 days after the mid-silkstage, i.e. the point when 50% of the plants within a given row hadreached the R1 (silking) growth stage, by injecting 5×105 Colletotrichumgraminicola spores suspended in 1 mL of distilled water. Thirty daysafter inoculation, the severity of anthracnose stalk rot in plants wasvisually assessed by splitting each stalk longitudinally to expose thepith. Each pith was examined to determine 1) the total number ofinternodes that displayed visual legions characteristic of the disease,and 2) the total number of internodes wherein visual legions hadinfected >75% of the tissue within the internode, as summarized in Table2. These two numbers were then summed into a disease score phenotype foreach plant, with scores of 10 converted to 9 to fit a scale ranging from1 (highly resistant) to 9 (highly susceptible). Twelve to fourteenplants were ranged per row and the space between each row was 0.80 m infield. The individual plant scores of each row were then averaged andthe average was reported as a final score for the row. Two populations(CV820914/CV391950 and CV094802/I294213*2) were measured in two fieldreplicates and one population (I283669*2/CV594360) was measured in threefield replicates for ASR resistance at different research sites usingmethods described in the art and the rating scale in Table 2.

Phenotype Analysis

After statistical procedures for phenotype quality control, a mixedmodel was run to estimate the variance components and to compute theheritability for ASR resistance. The heritability was 0.40 for inbredper se and 0.60 for hybrid for the population CV820914/CV391950, 0.46for inbred per se for the population CV094802/I294213*2, and 0.30 forinbred per se for the population I283669*2/CV594360.

TABLE 2 Rating Scale of relative ASR infection resistance phenotypes No.No. Internodes Internodes >75% Infected Infected Score Rating 1 0 1Highly resistant 1 1 2 Highly resistant 2 1 3 Resistant 2 2 4 Resistant3 2 5 Intermediate 3 3 6 Susceptible 4 3 7 Susceptible 4 4 8 Highlysusceptible 5 4 9 Highly susceptible 5 4 or 5 9 Highly susceptible

Primers and Probes Useful for Detecting ASR Resistance Genotypes

These plants were then genotyped using SNP markers that collectivelyspanned each chromosome in the maize genome. Loci that were monomorphicin the subject populations were eliminated from further analysis.

The primer sequences for amplifying exemplary SNP marker loci linked toASR-6.01 QTL and the probes used to genotype the corresponding SNPsequences are provided in Table 3. One of skill in the art willrecognize that sequences to either side of the given primers can be usedin place of the given primers, so long as the primers can amplify aregion that includes the allele to be detected. The precise probe to beused for detection can vary, e.g., any probe that can identify theregion of a marker amplicon to be detected can be substituted for thoseprobes exemplified herein. Also, configuration of the amplificationprimers and detection probes can, of course, vary. Thus, the inventionis not limited to the primers, probes, or marker sequences specificallyrecited herein.

TABLE 3 Primers and probes used for detecting SNPs linked to ASR-6.01 infield studies SEQ ID NO. SEQ SNP Fwd Rev ID NO. Pos. Primer Primer Probe1 Probe 2 1 433 11 21 31 41 2 381 12 22 32 42 3 221 13 23 33 43 4 130 1424 34 44 5 373 15 25 35 45 6 301 16 26 36 46 7 463 17 27 37 47 8 618 1828 38 48 9 148 19 29 39 49 10 469 20 30 40 50

Illustrative ASR resistance marker DNA sequences SEQ ID NOs: 1 can beamplified using the primers described in Table 3 as SEQ ID NOs: 11(forward primer) and 21 (reverse primer), and detected with probes asSEQ ID NOs: 31 (probe 1) and 41 (probe 2).

Marker-Trait Association Study

Marker-trait association studies were performed using both single-markeranalysis (SMA) and CIM. For each marker, the thresholds of Likelihoodratio between full and null models for CIM were based on 1000 randompermutation tests and the thresholds (p-value) for SMA were based on10,000 random permutation tests (Churchill and Doerge 1994). The CIManalysis revealed a strong QTL associated with ASR resistance onchromosome 6. The QTL was confirmed in multiple genetic backgrounds andmulti-year phenotypes for both inbred per se and hybrid populations. TheQTL peaks from these three populations were located on chromosome 6within 54 to 62 cMon the Monsanto's internal consensus genetic map asshown in Table 4. Combining the data from three mapping populations, theinterval for this QTL was 48.8-67.9 cM. This QTL is designated as“ASR-6.01”. The QTL effect for one copy of favorable allele was 0.75rating score and 1.5 rating score for homozygotes on average. Thephenotypic variance explained (R2) by this QTL was 24%.

TABLE 4 Summary of the CIM analysis from field studies A, B, and C.Field Population Resistant QTL P- QTL Total Study Mapping PopulationType #Mk Parent Chr Peak Left Right val Additive R² R² ACV820914/CV391950 Inbred 146 CV391950 6 53.9 48.8 65.1 0.01 0.75 0.240.48 A CV820914/CV391950 Hybrid 146 CV391950 6 54.9 49.9 67.9 0.01 1.160.55 0.66 B CV094802/I294213*2 Inbred 194 CV094802 6 55.7 49.7 57.5 0.010.77 0.11 0.35 C I283669*2/CV594360 Inbred 140 CV594360 6 62.5 56.8 66.60.01 0.61 0.11 0.22 *P-value is based on 1,000 permutation tests

Each row provides field study ID, mapping population, population type,number of markers used, resistant parent, chromosome position, the peakof the Likelihood ratio corresponds to ASR resistance, QTL intervalwhere left and right flanking positions are shown, additive effect,phenotypic variance of individual QTL R² and total R².

Table 5 lists the effect estimates on ASR resistance phenotype ratingsassociated with each marker (SEQ ID NO) measured by single markerassociation (SMA) analysis in field study A, B and C. Each row providesthe SEQ ID NO of the marker, and genetic map loci are represented in cM,with position zero being the first (most distal) marker known at thebeginning of the chromosome on the internal consensus genetic map,mapping population, Genetic source of favorable allele, favorableallele, unfavorable allele, F statistical value and the estimated effectthat the marker polymorphism had on the ASR phenotype. The statisticalsignificance (p-value) of the association between the marker and the ASRresistance rating in each case was p-value ≤0.01 on 10,000 permutationtests.

TABLE 5 Statistical associations of markers associated with ASR-6.01 infield studies A, B and C. Genetic Source of Permutation Single SEQ MONFavorable Favorable Unfavorable testing Allele ID NO. Map cM MappingPopulation Allele allele allele Fstat Probability Effect 1 49.7I283669*2/CV594360 CV594360 A T 14.1 0.01 0.51 2 62.5 I283669*2/CV594360CV594360 T C 23.4 0.001 0.63 3 49.6 CV820914/CV391950 CV391950 C T 19.40.001 0.54 4 53.9 CV820914/CV391950 CV391950 A G 45.3 0.001 0.77 5 63.1CV820914/CV391950 CV391950 T C 23.6 0.001 0.59 6 68.3 CV820914/CV391950CV391950 G A 21.2 0.001 0.56 7 49.7 CV094802/I294213*2 CV094802 T C 17.20.005 0.76 8 58.6 CV094802/I294213*2 CV094802 C G 17.6 0.005 0.78 9 69CV094802/I294213*2 CV094802 G A 19.4 0.001 0.81 *P-value is based on10,000 permutation tests

For example, SEQ ID NO: 3 was associated with a 0.54 change in ASRresistance rating by one copy of the favorable allele. SEQ ID NO: 5 wasassociated with a 0.59 change in ASR resistance rating by one copy ofthe favorable allele. ASR resistance ratings were generated using themethods described in Example 1.

Example 2. Field Study D

180 BC1F4 plants were derived from BC1F3 CV094802/I294213*2 as shown inTable 6. Corn plants were inoculated as described in Example 1 and thenmeasured for ASR resistance at research site using methods described inthe art and the rating scale in Table 2. These plants were genotypedusing SNP markers that collectively spanned each chromosome in the maizegenome. To note, the SNP markers used in this field study overlapped yetvaried from the SNP markers used in the prior field studies.

TABLE 6 Mapping populations of field study D Field Resistant SusceptiblePopulation Number Study Mapping Population Parent parent Type Gender ofLines D CV094802/I294213*2 CV094802 I294213 BC1F4 F 180

Marker-Trait Association Study

Marker-trait association studies were performed using both SMA and CIM.For each marker, the thresholds of Likelihood ratio between full andnull models for CIM were based on 1000 random permutation tests and thethresholds (p-value) for SMA were based on 10,000 random permutationtests (Churchill and Doerge 1994). The CIM analysis from field study Dconfirmed the QTL region associated with ASR resistance in Example 1.The QTL peak was mapped to 58.9 cM on chromosome 6 on theinternally-derived genetic map as shown in Table 7. The QTL interval was52.5-66.3 cM. The phenotypic variance explained (R²) by this QTL was15%.

TABLE 7 Summary of the CIM analysis from field study D Field PopulationResistant QTL P- QTL Total Study Type #Mk Parent Chr Peak Left Rightvalue Additive R² R² D Inbred 178 CV094802 6 58.9 52.5 66.3 0.01 0.540.15 0.38 *P-value is based on 1,000 permutation tests

Table 7 provides the population type, number of markers used, resistantparent, chromosome location, the peak of the Likelihood ratiocorresponds to ASR resistance, QTL interval where left and rightflanking positions are shown, additive effect, phenotypic variance ofindividual QTL or Total (R²).

Table 8 lists the effect estimates on ASR resistance phenotype ratingsof each marker (SEQ ID NO) linked to ASR-6.01 measured by single markerassociation (SMA) analysis from field study D. Each row provides the SEQID NO of the marker, genetic map loci are represented in cM, withposition zero being the first (most distal) marker known at thebeginning of the chromosome on the internal consensus genetic map,mapping population, genetic source of favorable allele, favorableallele, unfavorable allele, F statistical value and the estimated effectthat the marker polymorphism had on the ASR phenotype. The statisticalsignificance (p-value) of the association between the marker and the ASRresistance rating in each case was p-value ≤0.01 on 10,000 permutationtests.

TABLE 8 Estimate effects of markers associated with ASR-6.01 from fieldstudy D. Genetic Source of Permutation Single SEQ MON FavorableFavorable Unfavorable testing Allele ID NO. Map cM Mapping PopulationAllele allele allele Fstat Probability Effect 10 49.5 CV094802/I294213*2CV094802 A G 23.81 0.001 0.584 4 53.9 A G 28.18 0.001 0.618 2 62.5 T C29.14 0.001 0.625 *P-value is based on 10,000 permutation tests

For example, SEQ ID NO: 10 was associated with a 0.584 change in ASRresistance rating by one copy of the favorable allele. SEQ ID NO: 4 wasassociated with a 0.618 change in ASR resistance rating by one copy ofthe favorable allele. ASR resistance ratings were generated using themethods described in Example 1.

Example 3. First Round of Fine-Mapping of ASR Resistance QTL onChromosome 6

In order to obtain additional recombinants, CV820914/CV391950 derived F1lines were selected, self-crossed, and harvested. The resultingsegregating F2 kernels were chipped and genotyped with 8 SNP markerswithin 51-65 cM on Monsanto's internal consensus genetic map (Table 9).

TABLE 9 Primers and probes used for detecting SNPs linked to ASR-6.01 infirst round fine-mapping SEQ ID NO. SEQ SNP Fwd Rev ID NO. Pos. PrimerPrimer Probe 1 Probe 2 51 118 57 63 69 75 52 74 58 64 70 76 53 89 59 6571 77 8 618 18 28 38 48 54 101 60 66 72 78 55 291 61 67 73 79 5 373 1525 35 45 56 100 62 68 74 80

Kernels were bulked based on their haplotypes within this QTL region.These bulks (10-20 plants) were then planted and subsequently screenedfor ASR resistance via progeny test in the greenhouse. In Table 10, greycells indicated that the bulk carried the same alleles as thesusceptible inbred line at the specific SNP position. White cellsindicated that the bulk carried at least one copy of the same allele asthe resistant inbred line at the specific SNP position. Lines which werehomozygous or heterozygous for the resistant allele were groupedtogether in this analysis. Bulk “b” shared the same candidate QTL regionas the susceptible inbred line, CV820914. The individual plant scores ofeach bulk were averaged. The average was reported as a final score forthe bulk and then compared with that of bulk “b”. Bulk “f”, “g”, “h”,“i”, “j”, “p” and “o” displayed significantly reduced ASR severity(highlighted by black box, p-value <0.05). Based on the common SNP(highlighted by black oval) of these bulks, SEQ ID NO: 53 was identifiedas the peak marker. The two closest markers flanking SEQ ID NO: 53 areSEQ ID NO: 52 and SEQ ID NO: 8.

For example, bulk “f” shared at least one same allele as the resistantinbred line (CV391950) at the SNP positions represented by SEQ ID NO:51, SEQ ID NO: 52 and SEQ ID NO: 53 (highlighted by white cells). Bulk“f” shared the same alleles as the susceptible inbred line (CV820914) atthe SNP positions represented by SEQ ID NO: 8, SEQ ID NO: 54, SEQ ID NO:55, SEQ ID NO: 5 and SEQ ID NO: 56 (highlighted by grey cells).

Similar experiments were also conducted on BC1F2 kernels derived fromCV295879/CV391950. Bulk “f”, “g”, “i”, “j”, “p” and “o” displayedsignificantly reduced ASR severity (highlighted by black box, p-value<=0.05) compared with the bulk “b”. Among these resistant bulks, thesame common SNP (SEQ ID NO: 53, highlighted by black oval) wasidentified as the peak marker, as shown in Table 11.

The QTL region associated with ASR resistance was fine-mapped to53.9-58.6 cM on chromosome 6 based on the internally-derived genetic map(Table 12).

TABLE 12 Summary of first round fine-mapping results SEQ ID NO. MON MapcM Marker Profile 52 53.9 Left flanking marker 53 56.8 QTL peak 8 58.6Right flanking marker

Example 4. Second Round Fine-Mapping of ASR Resistance QTL on Chromosome6

In order to further fine-map the QTL region, CV005260/CV391950 derivedF2 lines were selected, self-crossed, and harvested. CV005260 was thesusceptible inbred line and CV391950 was the resistant inbred line. Theresulting segregating F3 kernels were chipped and genotyped with 6 SNPmarkers within 54-59 cM on the internally-derived genetic map (Table13). Primer and probe synthesis is within the skill of the art once theSNP position in the corn genome is provided. One of skill in the artwill also immediately recognize that other sequences to either side ofthe given primers can be used in place of the given primers, so long asthe primers can amplify a region that includes the allele to bedetected. Further, it will be appreciated that the precise probe to beused for detection can vary, e.g., any probe that can identify theregion of a marker amplicon to be detected can be substituted for thoseexamples provided herein. Also, configuration of the amplificationprimers and detection probes can, of course, vary. Thus, the inventionis not limited to the primers, probes, or marker sequences specificallyrecited herein.

TABLE 13 Primers and probes used for detecting SNPs linked to ASR- 6.01in second round fine-mapping from CV005260/CV391950 SEQ ID NO. SEQ SNPFwd Rev ID NO. Pos. Primer Primer Probe 1 Probe 2 4 130 14 24 34 44 81101 84 87 90 93 82 101 85 88 91 94 53 89 59 65 71 77 83 101 86 89 92 958 618 18 28 38 48

Kernels were bulked based on their haplotypes within this QTL region.These bulks (9-19 plants/bulk) were then planted and subsequentlyscreened for ASR resistance via progeny test in the greenhouse. In Table14, grey cells indicated that the bulk shared the same alleles as thesusceptible inbred line at the specific SNP positions. White cellsindicated that the bulk shared the same alleles as the resistant inbredline at the specific SNP positions. The individual plant scores of eachbulk were averaged and the average was reported as a final score for thebulk. Bulk “i”, “j”, “k”, “1” and “m” displayed significantly reducedASR severity with mean values less than 2 (highlighted by black box).Based on the common SNP (highlighted by black oval) of these bulks, SEQID NO: 83 was identified as the peak marker.

For example, bulk “a” shared the same alleles as the resistant inbredline, CV391950, at the candidate QTL region (highlighted by whitecells); bulk “b” shared the same alleles as the susceptible inbred line,CV005260, at the candidate QTL region (highlighted by grey cells).

The two closest markers flanking SEQ ID NO: 83 are SEQ ID NO: 53 and SEQID NO: 8. The QTL region associated with ASR resistance was furtherfine-mapped to 56.8-58.6 cM on chromosome 6 based on theinternally-derived genetic map (Table 15).

TABLE 15 Summary of second round fine-mapping results SEQ ID NO. MON MapcM Marker Profile 53 56.8 Left flanking marker 83 57.1 QTL peak 8 58.6Right flanking marker

Example 5. Further Fine-Mapping Using Genotype-by-Sequencing (GBS)Method

SNP markers were specifically designed for CV005260/CV391950-derivedplants via genotype-by-sequencing method within ASR-6.01 interval. Onehundred and twenty-seven BC1F2 inbred plants were genotyped and measuredfor ASR resistance. SMA analysis identified the top 11 SNP markersassociated with ASR resistance. Each row in Table 16 provides the SEQ IDNO of the marker, genetic map positions of the marker, SNP position,favorable allele, unfavorable allele, marker effect, p-value andphenotypic variance (R²) of the marker. Genetic map loci are representedin cM, with position zero being the first (most distal) marker known atthe beginning of the chromosome on Monsanto's internal consensus geneticmap.

TABLE 16 Further Fine-Mapping of ASR-6.01 via GBS Method SEQ MON SNPFavorable Unfavorable Marker ID NO. Map cM position allele allele Effectp-value R² 96 57.1 101 C T 1 0.0001 0.12 97 58 151 T C 0.94 0.0007 0.0998 58 151 T C 0.97 0.0002 0.11 99 58 151 T G 0.98 0.0001 0.11 100 58 151A G 0.98 0.0001 0.11 101 58 151 T G 0.98 0.0001 0.11 102 58 151 C A 0.980.0001 0.11 103 58 151 C T 0.97 0.0002 0.11 104 58.2 151 A G 1.93 0.00030.1 105 58.2 151 T A 0.75 0.0006 0.09 106 58.6 151 A G 0.71 0.0002 0.1

Chromosome intervals according to the invention and comprising markersclosely linked to the ASR-6.01 QTL are disclosed in Table 17. Geneticmap loci are represented in cM, with position zero being the first (mostdistal) marker known at the beginning of the chromosome on both aninternal consensus genetic map (MON) and the Neighbors 2008 maizegenomic map (IBM2008), which is freely available to the public from theMaize GDB website and commonly used by those skilled in the art. Alsodisclosed in Table 17 are the physical locations of loci as they arereported on the B73 RefGen_v2 sequence public assembly by the ArizonaGenomics Institute, available on the internet.

TABLE 17 Genetic and physical map positions of markers and chromosomeintervals associated with ASR-6.01. Relative Genetic Map Position† MONIBM2008 Physical Map Position†† Marker/Locus Map cM Map IcM Contig ChrStart Chr End IDP7601 38.2 200.7 AC204522.4 107366978 107368801 IDP6238.9 206.2 AC208541.3 107736319 107737212 l11 43.1 216 — — — IDP809044.6 219.5 AC201909.4 109532609 109539045 umc2006 48.7 228.9 — — —IDP8231 49.1 229.9 AC206946.2 113039145 113042824 umc248b 49.1 230 — — —SEQ ID NO: 10 49.5 231.1 — — — pco136292 49.6 231.3 — — — SEQ ID NO: 349.6 231.3 — — — IDP6025 49.7 231.5 AC212465.3 115308486 115310063IDP6010 49.7 231.5 AC209367.3 116123817 116124980 SEQ ID NO: 1 49.7231.6 — — — SEQ ID NO: 7 49.7 231.6 — — — agrr118a 50.1 232.6 — — —umc180(pep) 50.2 232.9 — — — SEQ ID NO: 51 50.7 234.3 — — — gpm74 51.1235.3 — — — TIDP3136 52.5 239.1 AC209629.2 118788679 118790325 AY10705353.7 242.3 — — — SEQ ID NO: 4 53.9 242.7 — — — SEQ ID NO: 52 53.9 242.7— — — IDP1699 54.1 243.3 AC197533.3 120878064 120883418 pdi7 54.9 245.3— — — gpm869 55.3 246.2 — — — SEQ ID NO: 81 55.9 247.6 — — — ufg11 56.1248.1 — — — SEQ ID NO: 82 56.3 251.3 — — — umc1250 56.6 254.5 AC194965.4127445565 127446395 SEQ ID NO: 53 56.8 255.2 — — — TIDP3356 57 255.9AC203836.3 128061342 128063001 SEQ ID NO: 83 57.1 256.3 — — — SEQ ID NO:96 57.1 256.3 — — — csu382a(cld) 57.3 257.1 — — — IDP2409 57.8 258.9AC189055.3 129901107 129902196 SEQ ID NO: 97 58 259.6 — — — SEQ ID NO:98 58 259.6 — — — SEQ ID NO: 99 58 259.6 — — — SEQ ID NO: 100 58 259.6 —— — SEQ ID NO: 101 58 259.6 — — — SEQ ID NO: 102 58 259.6 — — — SEQ IDNO: 103 58 259.6 — — — SEQ ID NO: 104 58.2 260.4 — — — SEQ ID NO: 10558.2 260.4 — — — SEQ ID NO: 106 58.6 262.3 — — — PCO146525 58.6 262.3 —— — SEQ ID NO: 8 58.6 262.3 — — — csu225 58.7 262.8 — — — bnl3.03 58.9264 — — — AI665560 61.1 273.2 AC195903.3 132064031 132064497 SEQ ID NO:54 61.5 274.4 — — — pzb00414 61.7 275.2 — — — umc2141 62.5 295.4AC204295.3 134846202 134846710 SEQ ID NO: 55 62.5 295.4 — — — SEQ ID NO:2 62.5 295.4 — — — AY110435 62.8 296.3 AC200260.3 137078584 137079677elfa5 63.1 297.1 — — — SEQ ID NO: 5 63.1 297.1 — — — umc1379 63.1 297.1AC214298.3 138536487 138537038 bnl15.37a 64.2 303 — — — SEQ ID NO: 5664.3 303.4 — — — pza02478 64.3 303.5 — — — IDP3886 65 306 AC205030.3141412746 141415456 cl39957_1 65.6 308 — — — mmc0241 66.2 312.8 — — —dup400(pac) 66.4 313.4 — — — jpsb107b 66.6 314 — — — chs562 67.9 317.5 —— — gpm709b 68.3 318.6 — — — SEQ ID NO: 6 68.3 318.7 — — — umc2321 68.4319 AC208555.3 147911766 147912501 bnlg1702 69 320.7 — — — SEQ ID NO: 969 320.7 — — — csu158b(eno) 69.2 321.6 — — — gpm426b 77.9 350.6 — — —†cM = centiMorgans, IcM = map units of the IBM2 2008 Neighbors GeneticMap. ††Arizona Genomics Institute B73 RefGen_v2 sequence. * Exactcoordinates not known. Coordinates can be estimated based on nearestflanking loci with known coordinates.

In Table 17, “IcM” refers to the map units of the IBM2 2008 NeighborsGenetic Map, which was generated with an intermated recombinant inbredpopulation (syn 4) that resulted in approximately a four-fold increasein the number of meiosies as compared to the typical recombinationexperiment that is used to generate centiMorgan (cM) distances (Lee etal., 2002, Plant Mol Biol 48:453 and the Maize Genetics and GenomicsDatabase). “cM” refers to the classical definition of a centimorganwherein one cM is equal to a 1% chance that a trait at one genetic locuswill be separated from a trait at another locus due to crossing over ina single generation (meaning the traits cosegregate 99% of the timeduring meiosis), and this definition is used herein to delineate maplocations pertaining to this invention. Any markers within theidentified region, including those disclosed herein or publicly known,could be used for detection of and selection for ASR resistance inaccordance with the methods of the present invention.

Example 6. Annotated Genes within ASR-6.01

Table 18 lists annotated coding sequences within ASR-6.01 region. Eachrow provides gene ID, gene annotation, chromosome location, geneticposition on Monsanto internal consensus map and physical position basedon Arizona Genomics Institute B73 RefGen_v2 sequence which is publiclyavailable. Transgenic maize resistant to anthracnose stalk rot can becreated using these annotated genes as described in the specification.

TABLE 18 Annotated coding sequences within ASR-6.01 region. MON MapPhysical Map Position bp †† Gene ID Annotation Chr cM † Start End 1Putative uncharacterized protein Sb10g013040 n = 2 Tax = AndropogoneaeRepID = C5Z298_SORBI (4e−20) 6 53.9 121384232 121384851 2 Delta zeinstorage protein n = 6 Tax = Zea mays RepID = C7AIP8_MAIZE (3e−22) 6 53.9121390345 121391190 3 S-layer domain protein n = 2 Tax = CyanotheceRepID = B7K1E8_CYAP8 (1e−19); GO_CC:GO:0009507, chloroplast# (3e−73) 653.9 121497243 121506368 4 ZCN11 n = 1 Tax = Zea mays RepID =A8WES3_MAIZE (5e−82); PBP: Phosphatidylethanolamine-binding protein 653.9 121537244 121538748 (9.1e−43); GO_BP:GO:0030154, celldifferentiation# (3e−34); GO_CC:GO:0005737, cytoplasm# (2e−46) 5 Proteinbinding protein n = 1 Tax = Zea mays RepID = B6SVM4_MAIZE (1e−103);GO_MF:GO:0046872, metal 6 53.9 121553074 121554257 ion binding#(1e−103); GO_BP:GO:0007229, integrin-mediated signaling pathway#(7e−25); GO_CC:GO:0005622, intracellular# (7e−35) 6 ELMOdomain-containing protein 2 n = 3 Tax = Zea mays RepID = B6T7G4_MAIZE(4e−23); 6 53.9 121554918 121556006 GO_BP:GO:0006909, IMP#phagocytosis#(4e−23); GO_CC:GO:0005856, cytoskeleton# (4e−23) 7 SMC4 protein n = 3Tax = Oryza sativa RepID = Q8L6H8_ORYSA (1e−104); GO_MF:GO:0005524, ATPbinding# (1e−104); 6 53.9 121556779 121559923 GO_BP:GO:0051276,chromosome organization# (1e−104); GO_CC:GO:0005694, chromosome#(1e−104) 8 Pollen-specific protein NTP303 n = 4 Tax = Zea mays RepID =B6U4I6_MAIZE (9e−60); Cu-oxidase: Multicopper 6 53.9 121566108 121567043oxidase (0.0056); GO_MF:GO:0016491, oxidoreductase activity# (9e−60);GO_BP:GO:0055114, oxidation reduction# (9e−60); GO_CC:GO:0005576,extracellular region# (5e−34) 9 AlphaSNBP(B)-like n = 3 Tax = Oryzasativa RepID = Q651K5_ORYSJ (1e−125); DFDF: DFDF motif (3.4e−07); 6 53.9121688996 121698144 FFD_TFG: FFD and TFG box motifs (1e−20) 10 HistonemRNA exonuclease 1 n = 2 Tax = Zea mays RepID = B6T5F9_MAIZE (0.0);Exonuc_X-T: Exonuclease 6 53.9 121757218 121777486 (8e−05);GO_MF:GO:0004527, exonuclease activity# (0.0); GO_BP:GO:0031125, rRNA3′-end processing# (2e−24); GO_CC:GO:0005622, intracellular# (2e−26) 11Aluminum-activated malate transporter-like n = 2 Tax = Oryza sativaRepID = Q6EPG5_ORYSJ (7e−11); GO_BP:GO:0010044, 6 53.9 121766949121767251 response to aluminum ion# (7e−11) 12 Putative polyprotein n =1 Tax = Zea mays RepID = Q8SA93_MAIZE (5e−45); Exo_endo_phos: 6 53.9121770395 121775874 Endonuclease/Exonuclease/phosphatase family (0.074);GO_MF:GO:0003964, RNA-directed DNA polymerase, group II intron encoded#(5e−45); GO_BP:GO:0015074, DNA integration# (5e−45); GO_CC:GO:0005634,nucleus# (5e−45) 13 DNA helicase homolog, putative n = 1 Tax = Musaacuminata RepID = Q1EPC6_MUSAC (1e−17); GO_MF:GO:0004386, 6 53.9121779361 121796764 helicase activity# (1e−17) 14 B-cellreceptor-associated protein 31-like containing protein n = 2 Tax =Andropogoneae RepID = B6TG43_MAIZE 6 53.9 121926217 121928807 (8e−20);GO_MF:GO:0004872, receptor activity# (8e−20); GO_BP:GO:0006886,intracellular protein transport# (2e−50); GO_CC:GO:0016021, integral tomembrane# (2e−50) 15 Reticulon n = 3 Tax = Andropogoneae RepID =B6TG01_MAIZE (1e−135); Reticulon: Reticulon (6.5e−58); 6 53.9 121943435121946102 GO_MF:GO:0003676, nucleic acid binding# (2e−97);GO_BP:GO:0006313, transposition, DNA-mediated# (8e−89);GO_CC:GO:0005783, IDA#endoplasmic reticulum# (1e−135) 16OSJNBa0029H02.21 protein n = 1 Tax = Oryza sativa RepID = Q7XT72_ORYSA(1e−09); Ribosomal_L23: 6 53.9 122086378 122086631 Ribosomal protein L23(0.0023); GO_MF:GO:0003735, structural constituent of ribosome# (2e−09);GO_BP:GO:0006412, translation# (2e−09); GO_CC:GO:0030529,ribonucleoprotein complex# (2e−09) 17 V-type proton ATPase 16 kDaproteolipid subunit c4 n = 52 Tax = Embryophyta RepID = VATL4_ARATH(2e−26); 6 53.9 122086738 122090981 GO_MF:GO:0015078, hydrogen iontransmembrane transporter activity# (2e−26); GO_BP:GO:0015992, protontransport# (2e−26); GO_CC:GO:0033179, proton-transporting V-type ATPase,V0 domain# (2e−26) 18 Putative uncharacterized protein n = 2 Tax = Zeamays RepID = B6U525_MAIZE (5e−10) 6 53.9 122093288 122093569 19Tesmin-like n = 2 Tax = Oryza sativa Japonica Group RepID = Q69WH4_ORYSJ(1e−145); CXC: Tesmin/TSO1- 6 53.9 122095245 122102827 like CXC domain(5.9e−16); CXC: Tesmin/TSO1-like CXC domain (1.4e−21); GO_MF:GO:0005515,protein binding# (1e−68); GO_BP:GO:0045449, regulation of transcription#(2e−74); GO_CC:GO:0031523, IDA#Myb complex# (8e−33) 20 Fiber proteinFb34 n = 2 Tax = Andropogoneae RepID = B4FVT6_MAIZE (7e−36); DUF1218:Protein of unknown 6 53.9 122116076 122116865 function (DUF1218)(2.2e−11) 21 Protein kinase family protein n = 2 Tax = Eumusa RepID =Q1EPA3_MUSAC (1e−146); Pkinase: Protein kinase 6 53.9 122247935122249667 domain (8.9e−36); Pkinase_Tyr: Protein tyrosine kinase(2.1e−36); APH: Phosphotransferase enzyme family (0.028);GO_MF:GO:0005524, ATP binding# (1e−179); GO_BP:GO:0006468, protein aminoacid phosphorylation# (1e−179); GO_CC:GO:0016459, myosin complex#(1e−179) 22 Putative Potential phospholipid-transporting ATPase 8 n = 2Tax = Oryza sativa RepID = Q67VX1_ORYSJ 6 53.9 122254921 122264424(0.0); E1-E2_ATPase: E1-E2 ATPase (9.6e−05); Hydrolase: haloaciddehalogenase-like hydrolase (0.0033); GO_MF:GO:0016820, hydrolaseactivity, acting on acid anhydrides, catalyzing transmembrane movementof substances# (0.0); GO_BP:GO:0016820, hydrolase activity, acting onacid anhydrides, catalyzing transmembrane movement of substances# (0.0);GO_CC:GO:0016021, integral to membrane# (0.0) 23 Retrotransposonprotein, putative, unclassified n = 1 Tax = Oryza sativa Japonica GroupRepID = 6 53.9 122304131 122304529 Q2QRU0_ORYSJ (1e−10); RVT_2: Reversetranscriptase (RNA-dependent DNA pol (0.0023); GO_MF:GO:0003676, nucleicacid binding# (2e−10); GO_BP:GO:0015074, DNA integration# (4e−10) 24Retrotransposon protein, putative, unclassified n = 2 Tax = Oryza sativaJaponica Group RepID = 6 53.9 122311854 122312338 Q10HG0_ORYSJ (3e−30);GO_MF:GO:0004523, ribonuclease H activity# (3e−30); GO_BP:GO:0006278,RNA-dependent DNA replication# (3e−30) 25Pyrophosphate-fructose-6-phosphate1-phosphotransferase alpha subunit(Fragment) n = 2 Tax = Saccharum 6 53.9 122322616 122325389 officinarumcomplex RepID = A1E380_SACSP (5e−24); GO_MF:GO:0003872,6-phosphofructokinase activity# (1e−24); GO_BP:GO:0006096, glycolysis#(1e−24); GO_CC:GO:0005945, 6-phosphofructokinase complex# (1e−24) 26Pho1-like protein n = 1 Tax = Populus trichocarpa RepID = B9HWP3_POPTR(0.0); SPX: SPX domain 6 53.9 122394825 122399345 (2.2e−26); EXS: EXSfamily (2.3e−144); GO_MF:GO:0004872, receptor activity# (0.0);GO_BP:GO:0004872, receptor activity# (0.0); GO_CC:GO:0016021, integralto membrane# (0.0) 27 PpPPR_77 protein n = 1 Tax = Physcomitrella patensRepID = Q5W963_PHYPA (1e−106); PPR: PPR 6 53.9 122404713 122406776repeat (0.26); PPR: PPR repeat (0.94); PPR: PPR repeat (0.0071); PPR:PPR repeat (3.6e−05); PPR: PPR repeat (6.1e−09); PPR: PPR repeat(0.057); PPR: PPR repeat (0.7); GO_MF:GO:0005488, binding# (4e−94);GO_CC:GO:0009536, plastid# (8e−99) 28 Putative uncharacterized protein9C20.7 n = 1 Tax = Zea mays RepID = Q5NKP3_MAIZE (1e−12) 6 53.9122421193 122421729 29 DEAD-box ATP-dependent RNA helicase 14 n = 4 Tax= Oryza sativa RepID = RH14_ORYSJ (2e−49); DEAD: 6 53.9 122524854122525657 DEAD/DEAH box helicase (0.087); GO_MF:GO:0016787, hydrolaseactivity# (2e−49); GO_BP:GO:0042254, ribosome biogenesis# (2e−49);GO_CC:GO:0005634, nucleus# (2e−49) 30 Putative uncharacterized protein n= 1 Tax = Zea mays RepID = B6SM34_MAIZE (2e−13) 6 53.9 122555403122555864 31 Pentatricopeptide repeat-containing protein, putative n = 1Tax = Ricinus communis RepID = B9SV96_RICCO 6 53.9 122582468 122585135(1e−173); PPR: PPR repeat (0.0007); PPR: PPR repeat (0.0031); PPR: PPRrepeat (9.6e−05); PPR: PPR repeat (5e−11); PPR: PPR repeat (0.0066);PPR: PPR repeat (0.036); PPR: PPR repeat (7e−10); PPR: PPR repeat(0.18); PPR: PPR repeat (8.7e−08); GO_MF:GO:0030528, transcriptionregulator activity# (0.0); GO_BP:GO:0045449, regulation oftranscription# (0.0); GO_CC:GO:0009536, plastid# (1e−173) 32 MADS-boxtranscription factor 31 n = 3 Tax = Andropogoneae RepID = B6TW19_MAIZE(6e−14); 6 53.9 122587039 122588859 GO_MF:GO:0043565, sequence-specificDNA binding# (6e−14); GO_BP:GO:0045449, regulation of transcription#(6e−14); GO_CC:GO:0005634, nucleus# (6e−14) 33 Probable calcium-bindingprotein CML30 n = 3 Tax = Oryza sativa RepID = CML30_ORYSJ (1e−54);efhand: EF 6 53.9 122664103 122665029 hand (0.0013); efhand: EF hand(0.006); efhand: EF hand (1.4e−06); efhand: EF hand (4e−08);GO_MF:GO:0005509, calcium ion storage activity# (1e−105);GO_BP:GO:0009409, IEP#response to cold# (5e−21); GO_CC:GO:0005737,cytoplasm# (5e−21) 34 Alkaline alpha galactosidase 2 n = 1 Tax = Zeamays RepID = Q575Z7_MAIZE (0.0); Raffinose_syn: Raffinose 6 53.9122727878 122731414 synthase or seed imbibition protein Sip1 (0);GO_MF:GO:0047274, galactinol-sucrose galactosyltransferase activity#(0.0); GO_BP:GO:0009409, IEP#response to cold# (0.0); GO_CC:GO:0009507,chloroplast# (0.0) 35 26S protease regulatory subunit 6A homolog n = 12Tax = Poaceae RepID = PRS6A_ORYSJ (2e−37); 6 53.9 122797640 122807850GO_MF:GO:0016765, transferase activity, transferring alkyl or aryl(other than methyl) groups# (2e−37); GO_BP:GO:0030163, protein catabolicprocess# (8e−36); GO_CC:GO:0005737, cytoplasm# (8e−36) 36 26S proteaseregulatory subunit 6A homolog n = 12 Tax = Poaceae RepID = PRS6A_ORYSJ(1e−135); AAA_2: 6 53.9 122836036 122838395 ATPase family associatedwith various (0.0093); AAA: ATPase family associated with variouscellular activities (AAA) (1.2e−89); AAA_3: ATPase family associatedwith various (0.011); AAA_5: ATPase family associated with various(0.00017); GO_MF:GO:0017111, nucleoside-triphosphatase activity#(1e−130); GO_BP:GO:0030163, protein catabolic process# (1e−130);GO_CC:GO:0005737, cytoplasm# (1e−130) 37 Putative uncharacterizedprotein n = 2 Tax = Zea mays RepID = B6TTZ4_MAIZE (0.0); Dev_Cell_Death:6 53.9 122889105 122892158 Development and cell death domain (1e−63);GO_MF:GO:0003677, DNA binding# (5e−33) 38 Longin-like n = 1 Tax =Medicago truncatula RepID = A4Q7K9_MEDTR (2e−77); Synaptobrevin:Synaptobrevin 6 53.9 122950872 122954048 (1.1e−39); GO_MF:GO:0005515,protein binding# (7e−45); GO_BP:GO:0016192, vesicle-mediated transport#(1e−104); GO_CC:GO:0016021, integral to membrane# (1e−104) 39 Clathrinassembly protein AP180 short form-like n = 2 Tax = Oryza sativa RepID =Q69SJ3_ORYSJ 6 53.95 122961662 122968145 (1e−169); ANTH: ANTH domain(5.7e−112); ENTH: ENTH domain (6.1e−05); GO_MF:GO:0030276, clathrinbinding# (0.0); GO_BP:GO:0048268, IDA#clathrin coat assembly# (0.0);GO_CC:GO:0030118, clathrin coat# (0.0) 40 Receptor serine-threonineprotein kinase, putative n = 1 Tax = Ricinus communis RepID =B9S1N3_RICCO 6 54 122994729 122996638 (1e−138); Pkinase: Protein kinasedomain (7.9e−31); Pkinase_Tyr: Protein tyrosine kinase (4.3e−31);GO_MF:GO:0005524, ATP binding# (0.0); GO_BP:GO:0006468, protein aminoacid phosphorylation# (0.0) 41 Cytochrome b-c1 complex subunit 8 n = 1Tax = Solanum tuberosum RepID = QCR8_SOLTU (2e−20); 6 54.1 123080089123083460 GO_MF:GO:0016491, oxidoreductase activity# (2e−16);GO_BP:GO:0022900, electron transport chain# (2e−20); GO_CC:GO:0070469,respiratory chain# (2e−20) 42 Putative iron/ascorbate-dependentoxidoreductase n = 1 Tax = Oryza sativa Japonica Group 6 54.1 123156144123157022 RepID = Q658E2_ORYSJ (7e−78); 2OG-FeII_Oxy: 2OG-Fe(II)oxygenase superfamily (1.1e−30); GO_MF:GO:0016491, oxidoreductaseactivity# (1e−80); GO_BP:GO:0055114, oxidation reduction# (1e−80) 43OSJNBa0029H02.21 protein n = 1 Tax = Oryza sativa RepID = Q7XT72_ORYSA(6e−22); Ribosomal_L23eN: 6 54.1 123201331 123203345 Ribosomal proteinL23, N-terminal domain (2.4e−18); GO_MF:GO:0003735, structuralconstituent of ribosome# (7e−22); GO_BP:GO:0006412, translation#(7e−22); GO_CC:GO:0030529, ribonucleoprotein complex# (7e−22) 44Putative uncharacterized protein Sb01g049710 n = 1 Tax = Sorghum bicolorRepID = C5X128_SORBI (1e−23) 6 54.1 123335141 123335768 45Pyrophosphate-energized vacuolar membrane proton pump n = 12 Tax =Poaceae RepID = AVP_HORVU (0.0); 6 54.1 123336536 123341421 H_PPase:Inorganic H+ pyrophosphatase (0); OPT: OPT oligopeptide transporterprotein (0.084); BCCT: BCCT family transporter (0.062);GO_MF:GO:0016787, hydrolase activity# (0.0); GO_BP:GO:0015992, protontransport# (0.0); GO_CC:GO:0016021, integral to membrane# (0.0) 46Heparanase-like protein 3 n = 3 Tax = Andropogoneae RepID = B6SXU7_MAIZE(0.0); Glyco_hydro_79n: 6 54.1 123344114 123346798 Glycosyl hydrolasefamily 79, N-terminal domain (2.4e−88); GO_MF:GO:0016798, hydrolaseactivity, acting on glycosyl bonds# (0.0); GO_BP:GO:0055085,transmembrane transport# (2e−50); GO_CC:GO:0016020, membrane# (0.0) 47Jp18 n = 1 Tax = Citrus trifoliata RepID = Q8H6R4_PONTR (3e−17) 6 54.1123370995 123372142 48 Nodulin-like protein n = 2 Tax = Oryza sativaRepID = Q8H613_ORYSJ (0.0); Nodulin-like: Nodulin-like 6 54.1 123372572123376208 (2.8e−117); MFS_1: Major Facilitator Superfamily (0.049);GO_MF:GO:0016798, hydrolase activity, acting on glycosyl bonds#(1e−153); GO_BP:GO:0055085, transmembrane transport# (0.0);GO_CC:GO:0016020, membrane# (1e−153) 49 Putative villin n = 1 Tax =Oryza sativa Japonica Group RepID = Q65XP6_ORYSJ (0.0); Gelsolin:Gelsolin 6 54.1 123383577 123388905 repeat (3.6e−11); Gelsolin: Gelsolinrepeat (0.00013); Gelsolin: Gelsolin repeat (5.4e−05); Gelsolin:Gelsolin repeat (2e−05); Gelsolin: Gelsolin repeat (0.064); Gelsolin:Gelsolin repeat (0.13); GO_MF:GO:0003779, actin binding# (0.0);GO_BP:GO:0007010, cytoskeleton organization# (0.0); GO_CC:GO:0015629,actin cytoskeleton# (0.0) 50 Tryptophan aminotransferase n = 2 Tax = Zeamays RepID = B5ATU2_MAIZE (0.0); Alliinase_C: Allinase, C- 6 54.2123432761 123435867 terminal domain (8.5e−198); Aminotran_1_2:Aminotransferase class I and II (0.00013); GO_MF:GO:0030170, pyridoxalphosphate binding# (0.0); GO_BP:GO:0080022, IMP#primary rootdevelopment# (7e−91); GO_CC:GO:0005737, cytoplasm# (2e−89) 51OSJNBa0053K19.25 protein n = 2 Tax = Oryza sativa RepID = Q7XPP8_ORYSJ(6e−26) 6 54.8 123669647 123669987 52 OSJNBa0053K19.25 protein n = 2 Tax= Oryza sativa RepID = Q7XPP8_ORYSJ (3e−46); DUF1682: Protein of 6 54.85123660711 123671949 unknown function (DUF1682) (0.0022);GO_CC:GO:0016021, integral to membrane# (2e−17) 53 OSJNBa0053K19.25protein n = 2 Tax = Oryza sativa RepID = Q7XPP8_ORYSJ (8e−18);GO_CC:GO:0005739, 6 54.9 123671681 123672761 mitochondrion# (2e−14) 542,3-bisphosphoglycerate-independent phosphoglycerate mutase n = 6 Tax =Poaceae RepID = PMGI_MAIZE 6 54.9 123673484 123674700 (3e−99); iPGM_N:BPG-independent PGAM N-terminus (iPGM (9.4e−16); GO_MF:GO:0046872, metalion binding# (3e−99); GO_BP:GO:0008152, metabolic process# (3e−99);GO_CC:GO:0005737, cytoplasm# (3e−99) 55 Cyclic nucleotide-gated ionchannel 2 (Fragment) n = 1 Tax = Hordeum vulgare subsp. vulgare 6 54.9123676263 123676490 RepID = Q4VDM4_HORVD (3e−15); GO_MF:GO:0005216, ionchannel activity# (5e−12); GO_BP:GO:0055085, transmembrane transport#(5e−12); GO_CC:GO:0016021, integral to membrane# (5e−12) 56 CENP-E likekinetochore protein n = 1 Tax = Zea mays RepID = B6SHI8_MAIZE (0.0);KIP1: KIP1-like protein 6 54.9 123677322 123687963 (2.6e−38); DUF2051:Double stranded RNA binding protein ((0.079); Pox_A_type_inc: ViralA-type inclusion protein repeat (57); Pox_A_type_inc: Viral A-typeinclusion protein repeat (77); Pox_A_type_inc: Viral A-type inclusionprotein repeat (0.79); Pox_A_type_inc: Viral A-type inclusion proteinrepeat (12); Cenp-F_leu_zip: Leucine-rich repeats of kinetochore p(0.053); Pox_A_type_inc: Viral A-type inclusion protein repeat (27);GO_MF:GO:0016301, kinase activity# (1e−132); GO_BP:GO:0016301, kinaseactivity# (1e−132); GO_CC:GO:0005886, plasma membrane# (1e−21) 57NHP2-like protein 1 n = 7 Tax = Andropogoneae RepID = B6TBE1_MAIZE(5e−24); GO_MF:GO:0005515, protein 6 54.9 123706476 123707839 binding#(5e−19); GO_BP:GO:0042254, ribosome biogenesis# (5e−24);GO_CC:GO:0030529, ribonucleoprotein complex# (5e−24) 58 Integralmembrane protein like n = 1 Tax = Zea mays RepID = B6SMU5_MAIZE(1e−139); UAA: UAA transporter 6 54.9 123729619 123734039 family(0.011); Nuc_sug_transp: Nucleotide-sugar transporter (0.022); DUF6:Integral membrane protein DUF6 (0.037); TPT: Triose-phosphateTransporter family (4.5e−49); GO_BP:GO:0009624, IEP#response tonematode# (5e−36); GO_CC:GO:0016021, integral to membrane# (1e−125) 59Aminotransferase y4uB n = 2 Tax = Andropogoneae RepID = B6T579_MAIZE(9e−27); GO_MF:GO:0030170, 6 54.9 123741189 123741689 pyridoxalphosphate binding# (9e−27); GO_BP:GO:0055114, oxidation reduction#(9e−12) 60 Aminotransferase y4uB n = 2 Tax = Andropogoneae RepID =B6T579_MAIZE (1e−62); GO_MF:GO:0030170, 6 54.9 123741701 123742472pyridoxal phosphate binding# (1e−62) 61 Putative uncharacterized proteinSb09g005000 n = 2 Tax = Andropogoneae RepID = C5Z118_SORBI (1e−111); 654.9 123748498 123751787 GO_BP:GO:0006979, response to oxidative stress#(2e−78) 62 Transcription factor BIM2 n = 3 Tax = Andropogoneae RepID =B6SVP6_MAIZE (7e−11); GO_MF:GO:0030528, 6 54.9 123753813 123761545transcription regulator activity# (7e−11); GO_BP:GO:0045449, regulationof transcription# (7e−11); GO_CC:GO:0005634, nucleus# (7e−11) 63 AP2domain-containing transcription factor n = 1 Tax = Populus trichocarpaRepID = B9GNL6_POPTR (4e−22); 6 54.9 123763332 123764175GO_MF:GO:0003700, transcription factor activity# (7e−23);GO_BP:GO:0045449, regulation of transcription# (7e−23);GO_CC:GO:0005634, nucleus# (7e−23) 64 Catalytic/hydrolase n = 2 Tax =Zea mays RepID = B6TIK5_MAIZE (1e−120); GO_MF:GO:0016787, hydrolase 654.9 123766758 123770464 activity# (1e−120); GO_BP:GO:0008152, metabolicprocess# (1e−120) 65 Germin-like protein n = 4 Tax = Andropogoneae RepID= Q6TM44_MAIZE (1e−117); Cupin_1: Cupin (3.6e−34); 6 54.9 123777183123778314 Cupin_2: Cupin domain (6.8e−07); GO_MF:GO:0046872, metal ionbinding# (8e−97); GO_BP:GO:0055114, oxidation reduction# (1e−71);GO_CC:GO:0048046, IDA#apoplast# (8e−97) 66 Ribulose bisphosphatecarboxylase large chain n = 4 Tax = BEP clade RepID = B8Y2Y5_FESAR(9e−63); 6 55.1 123912742 123913547 RuBisCO_large_N: Ribulosebisphosphate carboxylase large chain, N-terminal domain (1e−79);GO_MF:GO:0016984, ribulose-bisphosphate carboxylase activity# (4e−62);GO_BP:GO:0055114, oxidation reduction# (4e−62); GO_CC:GO:0009536,plastid# (4e−62) 67 Putative uncharacterized protein n = 1 Tax = Zeamays RepID = B8A1X9_MAIZE (3e−81) 6 55.1 123914152 123915257 68 AuxinEfflux Carrier family protein n = 1 Tax = Zea mays RepID = B6SVJ1_MAIZE(7e−29); 6 55.1 123949237 123949664 GO_BP:GO:0055085, transmembranetransport# (7e−29); GO_CC:GO:0016021, integral to membrane# (7e−29) 69Armadillo/beta-catenin-like repeat family protein n = 1 Tax = Zea maysRepID = B6U4A9_MAIZE (5e−32); 6 55.1 123952012 123952497GO_MF:GO:0005488, binding# (5e−32) 70 Kelch-like protein n = 3 Tax =Oryza sativa RepID = Q84S70_ORYSJ (7e−82); Dev_Cell_Death: Development 655.1 124019830 124042554 and cell death domain (7.1e−40); Kelch_1: Kelchmotif (0.095); Kelch_1: Kelch motif (1.3e−16); Kelch_2: Kelch motif(6.9e−05); Kelch_1: Kelch motif (4.2e−08); Kelch_2: Kelch motif (8.6);Kelch_1: Kelch motif (5.5e−11); Kelch_2: Kelch motif (0.00044); Kelch_1:Kelch motif (8.5e−14); Kelch_2: Kelch motif (1.5e−05); Kelch_1: Kelchmotif (4.4e−05); Kelch_2: Kelch motif (2.3); GO_MF:GO:0005515, proteinbinding# (5e−40); GO_BP:GO:0050807, IGI#regulation of synapseorganization# (7e−38); GO_CC:GO:0005575, cellular_component# (7e−38) 71Kelch-like protein n = 3 Tax = Oryza sativa RepID = Q84S70_ORYSJ(4e−44); Dev_Cell_Death: Development 6 55.1 124104544 124110586 and celldeath domain (2.8e−74); Kelch_2: Kelch motif (21); Kelch_1: Kelch motif(0.11); Kelch_1: Kelch motif (0.23); Kelch_2: Kelch motif (15); Kelch_1:Kelch motif (5.4e-12); Kelch_2: Kelch motif (2.3); Kelch_1: Kelch motif(1.2e−12); Kelch_2: Kelch motif (9.9e−06); Kelch_1: Kelch motif (0.004);Kelch_2: Kelch motif (2.4); GO_MF:GO:0005515, protein binding# (1e−26);GO_BP:GO:0046529, IGI#imaginal disc fusion, thorax closure# (6e−26);GO_CC:GO:0031463, IPI#Cul3-RING ubiquitin ligase complex# (6e−26) 72Erg28 like protein n = 4 Tax = Andropogoneae RepID = B6U3K5_MAIZE(9e−68); Erg28: Erg28 like protein 6 55.1 124135063 124140932 (1.6e−36);GO_BP:GO:0016126, IGI#sterol biosynthetic process# (1e−45);GO_CC:GO:0016021, integral to membrane# (9e−68) 73 Putativeuncharacterized protein Sb02g010980 n = 1 Tax = Sorghum bicolor RepID =C5X573_SORBI (6e−22) 6 55.1 124156120 124156572 74 DNA binding protein n= 1 Tax = Zea mays RepID = B6U8E0_MAIZE (1e−123); HLH: Helix-loop-helixDNA- 6 55.1 124197455 124198967 binding domain (3.5e−07);GO_MF:GO:0030528, transcription regulator activity# (1e−123);GO_BP:GO:0045449, regulation of transcription# (1e−123);GO_CC:GO:0005634, nucleus# (1e−123) 75 DNA helicase homolog, putative n= 1 Tax = Musa acuminata RepID = Q1EPC6_MUSAC (6e−10); 6 55.1 124205879124207211 GO_MF:GO:0004386, helicase activity# (6e−10) 76 RuBisCosubunit binding-protein beta subunit (Fragment) n = 1 Tax = Zea maysRepID = Q6B7Q9_MAIZE 6 55.1 124214939 124216073 (5e−62);GO_MF:GO:0005524, ATP binding# (7e−60); GO_BP:GO:0044267, cellularprotein metabolic process# (7e−60); GO_CC:GO:0009536, plastid# (7e−60)77 Ulp1 protease family, C-terminal catalytic domain containing proteinn = 2 Tax = Oryza sativa Japonica Group 6 55.1 124218673 124221589 RepID= Q109R5_ORYSJ (4e−37); Peptidase_C48: Ulp1 protease family, C-terminalcatalytic domain (4.9e−13); GO_MF:GO:0008234, cysteine-type peptidaseactivity# (1e−158); GO_BP:GO:0006508, proteolysis# (1e−158) 78Transposon protein, putative, CACTA, En/Spm sub-class n = 1 Tax = Oryzasativa Japonica Group 6 55.15 124153890 124156112 RepID = Q2QWY8_ORYSJ(1e−128); GO_MF:GO:0004803, transposase activity# (2e−57);GO_BP:GO:0006313, transposition, DNA-mediated# (2e−57) 79 Glutaryl-CoAdehydrogenase n = 1 Tax = Zea mays RepID = B6TNB5_MAIZE (0.0);Acyl-CoA_dh_N: Acyl-CoA 6 55.2 124146431 124151926 dehydrogenase,N-terminal domain (7e−30); Acyl-CoA_dh_M: Acyl-CoA dehydrogenase, middledomain (1.7e−22); Acyl-CoA_dh_1: Acyl-CoA dehydrogenase, C-terminal do(2.8e−25); Acyl-CoA_dh_2: Acyl-CoA dehydrogenase, C-terminal do(0.0014); GO_MF:GO:0050660, FAD binding# (0.0); GO_BP:GO:0055114,oxidation reduction# (0.0); GO_CC:GO:0009514, glyoxysome# (0.0) 80 PHDfinger transcription factor-like n = 2 Tax = Oryza sativa RepID =Q5JJV7_ORYSJ (1e−153); PHD: PHD-finger 6 55.3 124296440 124300947(1.8e−10); Acetyltransf_1: Acetyltransferase (GNAT) family (0.074);GO_MF:GO:0046872, metal ion binding# (1e−153); GO_BP:GO:0008152,metabolic process# (1e−153) 81 Growth inhibition anddifferentiation-related protein 88 n = 2 Tax = Andropogoneae RepID =B6TKU0_MAIZE 6 55.9 124363443 124366224 (1e−140); RNA_bind: RNA bindingdomain (0.027); GO_MF:GO:0046872, metal ion binding# (1e−140);GO_BP:GO:0006402, mRNA catabolic process# (1e−140); GO_CC:GO:0005737,cytoplasm# (1e−140) 82 Putative uncharacterized protein n = 1 Tax = Zeamays RepID = B4FS85_MAIZE (4e−14) 6 55.9 124471043 124472524 83 Putativeuncharacterized protein n = 3 Tax = Zea mays RepID = B6U549_MAIZE(4e−66) 6 55.9 124489396 124490221 84 Nucleolar complex protein 4 n = 1Tax = Zea mays RepID = B6SWX2_MAIZE (5e−43); DUF947: Domain of 6 55.9124525406 124526646 unknown function (DUF947) (0.018); CBF: CBF/Mak21family (2.7e−05); GO_MF:GO:0005515, protein binding# (1e−19);GO_BP:GO:0006364, rRNA processing# (4e−15); GO_CC:GO:0016020, membrane#(4e−34) 85 Fructose-6-phosphate-2-kinase/fructose-2,6-bisphosphatase n =4 Tax = Andropogoneae RepID = Q947C1_MAIZE 6 55.9 124531369 124550954(0.0); CBM_20: Starch binding domain (0.0071); 6PF2K:6-phosphofructo-2-kinase (2.6e−121); PGAM: Phosphoglycerate mutasefamily (1.6e−33); GO_MF:GO:0030246, carbohydrate binding# (0.0);GO_BP:GO:0016301, kinase activity# (0.0); GO_CC:GO:0043540,IDA#6-phosphofructo-2-kinase/fructose- 2,6-biphosphatase 1 complex#(8e−97) 86 RING-H2 finger protein ATL3F n = 2 Tax = Zea mays RepID =B6U6T9_MAIZE (2e−75); PHD: PHD-finger 6 55.9 124551008 124551883(0.066); zf-C3HC4: Zinc finger, C3HC4 type (RING finger) (2.8e−08);GO_MF:GO:0046872, metal ion binding# (2e−75); GO_BP:GO:0010200,IEP#response to chitin# (1e−15); GO_CC:GO:0016021, integral to membrane#(1e−15) 87 Putative CCR4-associated factor 1 n = 2 Tax = Oryza sativaRepID = Q5VPG5_ORYSJ (4e−72); CAF1: CAF1 6 55.9 124603474 124604545family ribonuclease (8.7e−63); GO_MF:GO:0003676, nucleic acid binding#(4e−72); GO_BP:GO:0045449, regulation of transcription# (3e−45);GO_CC:GO:0005634, nucleus# (4e−72) 88 Protein binding protein, putativen = 1 Tax = Ricinus communis RepID = B9S0N5_RICCO (2e−33); PHD: PHD- 656 124658046 124668099 finger (0.019); zf-C3HC4: Zinc finger, C3HC4 type(RING finger) (3.6e−07); GO_MF:GO:0046872, metal ion binding# (0.0) 89PHD finger protein n = 4 Tax = Andropogoneae RepID = B4FK95_MAIZE(1e−107); C1_3: C1-like domain 6 56.1 124705685 124710899 (0.091); PHD:PHD-finger (3.4e−10); GO_MF:GO:0046872, metal ion binding# (1e−107);GO_BP:GO:0046961, proton-transporting ATPase activity, rotationalmechanism# (2e−54); GO_CC:GO:0005634, nucleus# (4e−58) 90 Arginyl-tRNAsynthetase n = 2 Tax = Zea mays RepID = B4FMR1_MAIZE (0.0);Arg_tRNA_synt_N: Arginyl 6 56.2 124736591 124745075 tRNA synthetase Nterminal do (1e−18); tRNA-synt_1d: tRNA synthetases class I (R)(9.5e−117); DALR_1: DALR anticodon binding domain (3.7e−48);GO_MF:GO:0016874, ligase activity# (0.0); GO_BP:GO:0006420, arginyl-tRNAaminoacylation# (0.0); GO_CC:GO:0005737, cytoplasm# (0.0) 91Fasciclin-like arabinogalactan protein 8 n = 2 Tax = Zea mays RepID =B6SL10_MAIZE (1e−108); Fasciclin: 6 56.45 124803296 124805408 Fasciclindomain (4.3e−18); GO_CC:GO:0046658, anchored to plasma membrane# (2e−36)92 Arginine/serine-rich splicing factor, putative n = 1 Tax = Ricinuscommunis RepID = B9SGV2_RICCO (1e−39); 6 56.8 124861967 124863604zf-CCHC: Zinc knuckle (1.4e−05); zf-CCHC: Zinc knuckle (6.9e−06);GO_MF:GO:0046872, metal ion binding# (1e−39); GO_BP:GO:0008380, RNAsplicing# (2e−32) 93 Putative receptor-like protein kinase n = 2 Tax =Oryza sativa RepID = Q75IR9_ORYSJ (0.0); Pkinase: Protein 6 56.8124877371 124878770 kinase domain (1.9e−32); Pkinase_Tyr: Proteintyrosine kinase (2.3e−23); GO_MF:GO:0016301, kinase activity# (0.0);GO_BP:GO:0016301, kinase activity# (0.0); GO_CC:GO:0016021, integral tomembrane# (1e−141) 94 Aspartic proteinase oryzasin-1 n = 3 Tax = Zeamays RepID = B6TSQ9_MAIZE (3e−51); GO_MF:GO:0016787, 6 56.8 124956620124957402 hydrolase activity# (3e−51); GO_BP:GO:0006629, lipid metabolicprocess# (3e−51); GO_CC:GO:0005773, IDA#vacuole# (1e−41) 95 Peroxidase(Fragment) n = 6 Tax = Zea mays RepID = Q6RFK0_MAIZE (0.0); peroxidase:Peroxidase (2.4e−134); 6 56.8 125011395 125012945 GO_MF:GO:0046872,metal ion binding# (0.0); GO_BP:GO:0055114, oxidation reduction# (0.0);GO_CC:GO:0016021, integral to membrane# (2e−84) 96 Peroxidase (Fragment)n = 6 Tax = Zea mays RepID = Q6RFK0_MAIZE (1e−157); peroxidase:Peroxidase 6 56.8 125155203 125156888 (4.3e−134); GO_MF:GO:0046872,metal ion binding# (0.0); GO_BP:GO:0055114, oxidation reduction# (0.0);GO_CC:GO:0016021, integral to membrane# (5e−86) 97 Chloroplast RelAhomologue 2 n = 2 Tax = Oryza sativa Japonica Group RepID = Q9AYT4_ORYSJ(0.0); 6 56.8 125187331 125190275 RelA_SpoT: Region found in RelA/SpoTproteins (6.3e−46); efhand: EF hand (0.0012); efhand: EF hand (0.043);GO_MF:GO:0005509, calcium ion storage activity# (0.0); GO_BP:GO:0015969,guanosine tetraphosphate metabolic process# (0.0); GO_CC:GO:0009507,chloroplast# (1e−149) 98 Dihydrolipoyl dehydrogenase n = 3 Tax = PoaceaeRepID = B9FML1_ORYSJ (1e−61); Pyr_redox_2: Pyridine 6 56.8 125286546125310756 nucleotide-disulphide oxidored (0.0013); Pyr_redox: Pyridinenucleotide-disulphide oxidore (8.3e−16); GO_MF:GO:0050660, FAD binding#(1e−61); GO_BP:GO:0055114, oxidation reduction# (1e−61);GO_CC:GO:0005737, cytoplasm# (1e−61) 99 Retrotransposon protein,putative, unclassified n = 1 Tax = Oryza sativa Japonica Group RepID =Q2QSA6_ORYSJ 6 56.8 125334886 125345192 (1e−70); GO_MF:GO:0003677, DNAbinding# (1e−70); GO_BP:GO:0015074, DNA integration# (1e−70);GO_CC:GO:0005634, nucleus# (7e−67) 100 Alpha-L-fucosidase 2 n = 3 Tax =Zea mays RepID = B6TDT3_MAIZE (0.0); Lipase_GDSL: GDSL-like 6 56.8125454576 125459572 Lipase/Acylhydrolase (9.8e−76); GO_MF:GO:0016788,hydrolase activity, acting on ester bonds# (0.0); GO_BP:GO:0006629,lipid metabolic process# (0.0); GO_CC:GO:0005576, extracellular region#(8e−88) 101 Ubiquitin-protein ligase, putative n = 1 Tax = Ricinuscommunis RepID = B9RZW1_RICCO (1e−114); HECT: 6 56.8 125545211 125554393HECT-domain (ubiquitin-transferase) (4.5e-53); GO_MF:GO:0016881,acid-amino acid ligase activity# (1e−123); GO_BP:GO:0006464, proteinmodification process# (1e−123); GO_CC:GO:0005622, intracellular#(1e−123) 102 Ubiquitin-protein ligase, putative n = 1 Tax = Ricinuscommunis RepID = B9RZW1_RICCO (2e−32); 6 56.8 125612410 125613737GO_MF:GO:0016881, acid-amino acid ligase activity# (7e−46);GO_BP:GO:0006464, protein modification process# (7e−46);GO_CC:GO:0005622, intracellular# (7e−46) 103 Putative aldosereductase-related protein n = 1 Tax = Zea mays RepID = Q7FS90_MAIZE(2e−19); Kelch_1: 6 56.8 125628803 125629340 Kelch motif (1.6e−06);Kelch_2: Kelch motif (0.00097); GO_MF:GO:0005515, protein binding#(5e−46); GO_BP:GO:0055114, oxidation reduction# (2e−19) 104 Proteinphosphatase 2C 35 n = 2 Tax = Oryza sativa RepID = P2C35_ORYSJ (5e−40);GO_MF:GO:0046872, 6 56.8 125630885 125631928 metal ion binding# (5e−40);GO_BP:GO:0006952, defense response# (5e−40); GO_CC:GO:0016020, membrane#(5e−40) 105 Replication protein A 70 kDa DNA-binding subunit n = 3 Tax =Zea mays RepID = B6SL03_MAIZE (1e−141); 6 56.8 125632605 125635951Rep_fac-A_C: Replication factor-A C terminal domain (4e−66);GO_MF:GO:0003677, DNA binding# (1e−141); GO_BP:GO:0006260, DNAreplication# (1e−141); GO_CC:GO:0005634, nucleus# (1e−141) 106Retrotransposon protein, putative, Ty1-copia subclass n = 1 Tax = Oryzasativa Japonica Group 6 56.8 125879055 125879270 RepID = Q2R0F7_ORYSJ(8e−17); GO_MF:GO:0003677, DNA binding# (8e−17); GO_BP:GO:0015074, DNAintegration# (8e−17) 107 Gibberellin 2-oxidase n = 1 Tax = Zea maysRepID = B6U889_MAIZE (1e−139); 2OG-FeII_Oxy: 2OG-Fe(II) 6 56.8 125881243125886353 oxygenase superfamily (0.008); GO_MF:GO:0016491,oxidoreductase activity# (1e−139); GO_BP:GO:0055114, oxidationreduction# (1e−139); GO_CC:GO:0016020, membrane# (3e−42) 108 Putativegag-pol polyprotein n = 1 Tax = Oryza sativa Japonica Group RepID =Q6UUN3_ORYSJ (4e−33); 6 56.8 125909464 125910136 GO_MF:GO:0004190,penicillopepsin activity# (4e−33); GO_BP:GO:0015074, DNA integration#(4e−33); GO_CC:GO:0005634, nucleus# (9e−29) 109 ATP citrate lyaseb-subunit n = 3 Tax = Papilionoideae RepID = Q93YH3_LUPAL (6e−21);GO_MF:GO:0005524, 6 56.8 126000743 126001820 ATP binding# (6e−23);GO_BP:GO:0006085, NAS#acetyl-CoA biosynthetic process# (1e−19) 110Putative aspartic proteinase nepenthesin I n = 1 Tax = Oryza sativaJaponica Group RepID = Q69IP6_ORYSJ 6 56.8 126012128 126013728 (9e−45);Asp: Eukaryotic aspartyl protease (1.6e−05); GO_MF:GO:0004190,penicillopepsin activity# (9e−45); GO_BP:GO:0006508, proteolysis#(9e−45); GO_CC:GO:0005576, extracellular region# (4e−41) 111 Transposonprotein, putative, CACTA, En/Spm sub-class n = 1 Tax = Oryza sativaJaponica Group 6 56.8 126014170 126016932 RepID = Q10BK1_ORYSJ (5e−59);GO_MF:GO:0004803, transposase activity# (4e−92); GO_BP:GO:0006313,transposition, DNA-mediated# (4e−92); GO_CC:GO:0016020, membrane#(1e−33) 112 Putative uncharacterized protein n = 1 Tax = Zea mays RepID= C0PP88_MAIZE (3e−32) 6 56.8 126063923 126064225 113 Putativeuncharacterized protein Sb09g004490 n = 1 Tax = Sorghum bicolor RepID =C5Z0L6_SORBI (4e−18); 6 56.8 126090413 126091289 GO_MF:GO:0030528,transcription regulator activity# (7e−10); GO_BP:GO:0045449, regulationof transcription# (7e−10); GO_CC:GO:0005634, nucleus# (7e−10) 114Putative uncharacterized protein n = 1 Tax = Zea mays RepID =C4JC40_MAIZE (8e−11) 6 56.8 126126609 126126863 115 CID11 n = 3 Tax =Andropogoneae RepID = B4FZ16_MAIZE (1e−41); GO_MF:GO:0003676, nucleicacid 6 56.8 126130008 126133974 binding# (1e−41); GO_BP:GO:0006397, mRNAprocessing# (4e−20); GO_CC:GO:0005634, nucleus# (4e−20) 116 USP familyprotein n = 4 Tax = Andropogoneae RepID = B6TC12_MAIZE (4e−65); Usp:Universal stress protein 6 56.8 126146579 126149276 family (1.6e−20);GO_MF:GO:0016818, hydrolase activity, acting on acid anhydrides, inphosphorus-containing anhydrides# (1e−55); GO_BP:GO:0006950, response tostress# (4e−65); GO_CC:GO:0005634, nucleus# (1e−55) 117 DNA-directed RNApolymerase II 19 kDa polypeptide n = 2 Tax = Andropogoneae RepID =B4FXC7_MAIZE 6 56.8 126149278 126152151 (2e−98); RNA_pol_Rpb7_N: RNApolymerase Rpb7, N-terminal domain (6.2e−13); RNA_pol_Rbc25: RNApolymerase III subunit Rpc25 (0.059); GO_MF:GO:0003899, DNA-directed RNApolymerase III activity# (2e−98); GO_BP:GO:0006350, transcription#(2e−98); GO_CC:GO:0080137, IPI#DNA-directed RNA polymerase V complex#(1e−48) 118 Putative uncharacterized protein Sb10g005530 n = 1 Tax =Sorghum bicolor RepID = C5Z5H0_SORBI (6e−25) 6 56.8 126152259 126152825119 Pyrophosphate-energized vacuolar membrane proton pump n = 2 Tax =Andropogoneae RepID = B6UEE8_MAIZE 6 56.8 126196881 126200082 (0.0);H_PPase: Inorganic H+ pyrophosphatase (0); OPT: OPT oligopeptidetransporter protein (0.048); DUF540: Protein of unknown function(DUF540) (0.096); GO_MF:GO:0009678, hydrogen-translocatingpyrophosphatase activity# (0.0); GO_BP:GO:0015992, proton transport#(0.0); GO_CC:GO:0016020, membrane# (0.0) 120 Putative uncharacterizedprotein n = 1 Tax = Zea mays RepID = B6U5J9_MAIZE (2e−16) 6 56.8126286798 126287445 121 Putative polyprotein n = 1 Tax = Oryza sativaJaponica Group RepID = Q65XD2_ORYSJ (2e−16); 6 56.8 126290188 126291608GO_MF:GO:0004190, penicillopepsin activity# (2e−16); GO_BP:GO:0015074,DNA integration# (2e−16); GO_CC:GO:0005634, nucleus# (8e−16) 122Polyprotein n = 1 Tax = Oryza sativa Japonica Group RepID = Q8W150_ORYSJ(1e−30); GO_MF:GO:0003964, 6 56.8 126292009 126292632 RNA-directed DNApolymerase, group II intron encoded# (1e−30); GO_BP:GO:0015074, DNAintegration# (1e−30); GO_CC:GO:0005634, nucleus# (1e−30) 123 PutativeSMEK homolog 3 n = 2 Tax = Mus musculus RepID = SMEK3_MOUSE (8e−19);GO_MF:GO:0005488, 6 56.8 126343969 126349725 binding# (2e−47) 124Putative polyprotein n = 1 Tax = Zea mays RepID = Q8SA93_MAIZE (1e−160);DUF625: Protein of unknown 6 56.8 126358241 126418203 function (DUF625)(8.6e−57); GO_MF:GO:0003964, RNA-directed DNA polymerase, group IIintron encoded# (1e−160); GO_BP:GO:0015074, DNA integration# (1e−160);GO_CC:GO:0005634, nucleus# (1e−160) 125 OSJNBa0065O17.7 protein n = 1Tax = Oryza sativa Japonica Group RepID = Q7XPS4_ORYSJ (2e−40); 6 56.8126362377 126368009 GO_MF:GO:0003964, RNA-directed DNA polymerase, groupII intron encoded# (2e−40); GO_BP:GO:0015074, DNA integration# (2e−40)126 Tubulin gamma-1 chain n = 28 Tax = Embryophyta RepID = TBG1_ARATH(0.0); Tubulin: Tubulin/FtsZ family, 6 56.8 126579166 126583379 GTPasedomain (3.7e−94); Tubulin_C: Tubulin/FtsZ family, C-terminal domain(1.2e−58); GO_MF:GO:0005525, GTP binding# (0.0); GO_BP:GO:0051641,IMP#cellular localization# (0.0); GO_CC:GO:0043234, protein complex#(0.0) 127 Putative uncharacterized protein Sb01g009200 n = 3 Tax =Andropogoneae RepID = C5WLY8_SORBI (1e−26); 6 56.8 126648784 126649020GO_MF:GO:0008375, acetylglucosaminyltransferase activity# (1e−11);GO_CC:GO:0016020, membrane# (1e−11) 128 Chaperone protein dnaJ, putativen = 1 Tax = Ricinus communis RepID = B9RNG7_RICCO (1e−141); DnaJ: DnaJ 656.8 126678899 126686639 domain (1e−36); DnaJ_C: DnaJ C terminal region(1.6e−18); GO_MF:GO:0051082, unfolded protein binding# (1e−145);GO_BP:GO:0006457, protein folding# (1e−145); GO_CC:GO:0005886, plasmamembrane# (1e−139) 129 Putative uncharacterized protein Sb02g029480 n =2 Tax = Andropogoneae RepID = C5X5A3_SORBI (8e−23); 6 56.8 126761961126762272 GO_MF:GO:0005515, protein binding# (5e−16); GO_CC:GO:0005737,cytoplasm# (5e−16) 130 Protein disulfide isomerase n = 2 Tax =Andropogoneae RepID = Q5EUD6_MAIZE (0.0); Thioredoxin: 6 56.8 126779705126784442 Thioredoxin (2.5e−49); AhpC-TSA: AhpC/TSA family (0.034);Thioredoxin: Thioredoxin (2.9e−57); ERp29: Endoplasmic reticulum proteinERp29, C-te (3.3e−46); GO_MF:GO:0016853, isomerase activity# (0.0);GO_BP:GO:0045454, cell redox homeostasis# (0.0); GO_CC:GO:0005783,IDA#endoplasmic reticulum# (0.0) 131 Retrotransposon protein, putative,unclassified n = 2 Tax = Oryza sativa Japonica Group RepID =Q2QZV1_ORYSJ 6 56.8 126790130 126790810 (2e−26); GO_MF:GO:0003964,RNA-directed DNA polymerase, group II intron encoded# (2e−29);GO_BP:GO:0006278, RNA-dependent DNA replication# (2e−29);GO_CC:GO:0005634, nucleus# (4e−21) 132 Gamma-tubulin complex component,putative n = 1 Tax = Ricinus communis RepID = B9SAS5_RICCO (0.0); 6 56.8126812997 126837059 Spc97_Spc98: Spc97/Spc98 family (9.1e−144);GO_MF:GO:0005515, protein binding# (0.0); GO_BP:GO:0000226, microtubulecytoskeleton organization# (0.0); GO_CC:GO:0005815, microtubuleorganizing center# (0.0) 133 Putative AC transposase n = 1 Tax = Zeamays RepID = TRA1_MAIZE (0.0); zf-BED: BED zinc finger (4.4e−05); 6 56.8126822114 126824371 hATC: hAT family dimerisation domain (1.2e−38);GO_MF:GO:0046983, protein dimerization activity# (0.0);GO_BP:GO:0032196, transposition# (0.0) 134 Putative polyprotein n = 1Tax = Zea mays RepID = Q8SA93_MAIZE (2e−26); GO_MF:GO:0003964, RNA- 656.8 126826110 126831487 directed DNA polymerase, group II intronencoded# (2e−26); GO_BP:GO:0015074, DNA integration# (2e−26);GO_CC:GO:0005634, nucleus# (2e−26) 135 F6D8.18 protein n = 11 Tax =rosids RepID = Q9SSR2_ARATH (1e−22); GO_MF:GO:0008233, peptidaseactivity# 6 56.8 126872786 126877517 (1e−24); GO_BP:GO:0006508,proteolysis# (1e−24); GO_CC:GO:0016020, membrane# (1e−24) 136 Ulp1protease family, C-terminal catalytic domain containing protein n = 2Tax = Oryza sativa Japonica Group 6 56.8 126877369 126879300 RepID =Q109R5_ORYSJ (5e−34); Peptidase_C48: Ulp1 protease family, C-terminalcatalytic domain (5.8e−13); GO_MF:GO:0008234, cysteine-type peptidaseactivity# (1e−129); GO_BP:GO:0006508, proteolysis# (1e−129) 137 VIP2protein n = 1 Tax = Avena fatua RepID = Q9M4C5_AVEFA (1e−171); zf-C3HC4:Zinc finger, C3HC4 type 6 56.8 126957235 126967077 (RING finger)(4.8e−05); zf-RING-like: RING-like domain (0.095); GO_MF:GO:0046872,metal ion binding# (1e−171); GO_BP:GO:0004842, NAS#ubiquitin-proteinligase activity# (2e−43) 138 Putative ABI3-interacting protein 2 n = 1Tax = Oryza sativa Japonica Group RepID = Q6K486_ORYSJ (5e−09); 6 56.8126982783 126983154 GO_BP:GO:0000226, microtubule cytoskeletonorganization# (5e−09); GO_CC:GO:0005874, microtubule# (5e−09) 139 ATPbinding protein n = 2 Tax = Andropogoneae RepID = B6SXM5_MAIZE (0.0);Kinesin: Kinesin motor domain 6 56.8 127109946 127127395 (1.2e−125);GO_MF:GO:0005524, ATP binding# (0.0); GO_BP:GO:0007018,microtubule-based movement# (0.0); GO_CC:GO:0005874, microtubule# (0.0)140 Putative polyprotein n = 1 Tax = Zea mays RepID = Q8SA93_MAIZE(2e−62); RVT_1: Reverse transcriptase 6 56.8 127112084 127116226(RNA-dependent DN (0.00015); GO_MF:GO:0003964, RNA-directed DNApolymerase, group II intron encoded# (2e−62); GO_BP:GO:0015074, DNAintegration# (2e−62); GO_CC:GO:0005634, nucleus# (2e−62) 141 Putativepolyprotein n = 1 Tax = Zea mays RepID = Q8SA93_MAIZE (1e−28);GO_MF:GO:0003964, RNA- 6 56.8 127116846 127117349 directed DNApolymerase, group II intron encoded# (1e−28); GO_BP:GO:0006355,regulation of transcription, DNA-dependent# (1e−34); GO_CC:GO:0005634,nucleus# (1e−34) 142 Putative polyprotein n = 1 Tax = Zea mays RepID =Q8SA93_MAIZE (6e−62); GO_MF:GO:0003964, RNA- 6 56.8 127129438 127130136directed DNA polymerase, group II intron encoded# (6e−62);GO_BP:GO:0015074, DNA integration# (6e−62); GO_CC:GO:0005634, nucleus#(6e−62) 143 Serine/threonine-protein phosphatase n = 2 Tax =Andropogoneae RepID = C5Z0J0_SORBI (1e−175); Metallophos: 6 56.8127199789 127203947 Calcineurin-like phosphoesterase (1.3e−44);GO_MF:GO:0016787, hydrolase activity# (1e−163); GO_BP:GO:0004721,phosphoprotein phosphatase activity# (1e−163); GO_CC:GO:0016459, myosincomplex# (1e−128) 144 Putative uncharacterized protein n = 1 Tax = Zeamays RepID = B6UAL2_MAIZE (2e−23) 6 56.8 127262064 127265900 145Putative uncharacterized protein n = 1 Tax = Zea mays RepID =B6UAL2_MAIZE (3e−17) 6 56.8 127283052 127283372 146 HAT familydimerisation domain containing protein n = 1 Tax = Oryza sativa JaponicaGroup 6 56.8 127360096 127364045 RepID = Q8LNK9_ORYSJ (0.0); zf-BED: BEDzinc finger (9.6e−08); hATC: hAT family dimerisation domain (4.6e−39);GO_MF:GO:0046983, protein dimerization activity# (0.0);GO_BP:GO:0006468, protein amino acid phosphorylation# (3e−92) 147Ribosomal protein L18 n = 16 Tax = Poaceae RepID = Q5WMY3_ORYSJ (3e−99);Ribosomal_L18e: Eukaryotic 6 56.8 127372821 127376997 ribosomal proteinL18 (7.2e−127); GO_MF:GO:0003735, structural constituent of ribosome#(3e−99); GO_BP:GO:0006412, translation# (3e−99); GO_CC:GO:0030529,ribonucleoprotein complex# (3e−99) 148 Ethylene receptor protein n = 1Tax = Musa acuminata AAA Group RepID = A1IIY0_MUSAC (0.0); GAF: GAF 656.8 127377761 127381836 domain (1.6e−07); HisKA: His Kinase A(phosphoacceptor) domain (6.3e−20); HATPase_c: Histidine kinase-, DNAgyrase B-, and HSP90-like ATPase (4.6e−31); GO_MF:GO:0016772,transferase activity, transferring phosphorus-containing groups# (0.0);GO_BP:GO:0018106, peptidyl-histidine phosphorylation# (0.0);GO_CC:GO:0016020, membrane# (0.0) 149 Putative uncharacterized proteinSb09g004315 (Fragment) n = 1 Tax = Sorghum bicolor RepID = C5Z0J6_SORBI6 56.8 127445565 127447307 (1e−14); GO_MF:GO:0046872, metal ion binding#(2e−09); GO_BP:GO:0006355, regulation of transcription, DNA-dependent#(2e−09) 150 Importin subunit alpha-1b n = 5 Tax = Poaceae RepID =IMA1B_ORYSJ (0.0); IBB: Importin beta binding domain 6 56.8 127496705127501662 (8.7e−27); Arm: Armadillo/beta-catenin-like repeat (4); HEAT:HEAT repeat (30); Arm: Armadillo/beta-catenin- like repeat (5.1e−11);HEAT: HEAT repeat (5.1); Arm: Armadillo/beta-catenin-like repeat(1.3e−14); HEAT: HEAT repeat (8.1e−05); Arm: Armadillo/beta-catenin-likerepeat (4.5e−08); Arm: Armadillo/beta-catenin-like repeat (1.1e−06);HEAT: HEAT repeat (1.5); Arm: Armadillo/beta-catenin-like repeat(2.1e−10); HEAT: HEAT repeat (1.5); Arm: Armadillo/beta-catenin-likerepeat (8e−11); HEAT: HEAT repeat (9.3); Arm: Armadillo/beta-catenin-like repeat (1.7e−13); HEAT: HEAT repeat (30); Arm:Armadillo/beta-catenin-like repeat (4.5e−07); HEAT: HEAT repeat (37);GO_MF:GO:0008565, protein transporter activity# (0.0); GO_BP:GO:0015031,protein transport# (0.0); GO_CC:GO:0048471, ISS#perinuclear region ofcytoplasm# (0.0) 151 WRKY67-superfamily of TFs having WRKY and zincfinger domains n = 2 Tax = Zea mays 6 56.8 127590782 127592172 RepID =B6T4Y9_MAIZE (7e−51); FAR1: FAR1 family (0.0056); WRKY: WRKY DNA-bindingdomain (7.5e−36); GO_MF:GO:0043565, sequence-specific DNA binding#(4e−81); GO_BP:GO:0045449, regulation of transcription# (4e−81);GO_CC:GO:0005634, nucleus# (4e−81) 152 Putative uncharacterized proteinSb05g019580 n = 1 Tax = Sorghum bicolor RepID = C5Y395_SORBI (2e−10) 656.8 127598820 127599045 153 Putative uncharacterized protein n = 2 Tax= Zea mays RepID = B6SQA8_MAIZE (8e−56) 6 56.8 127633793 127634935 154H0103C06.4 protein n = 1 Tax = Oryza sativa RepID = Q259H8_ORYSA(4e−28); GO_MF:GO:0046983, protein 6 56.8 127659907 127661365dimerization activity# (3e−28) 155 Alpha-L-fucosidase 2 n = 2 Tax = Zeamays RepID = B6TLP8_MAIZE (2e−66); Lipase_GDSL: GDSL-like 6 56.8127663965 127666621 Lipase/Acylhydrolase (3.5e−07); Gp_dh_N:Glyceraldehyde 3-phosphate dehydrogenase, (1.3e−07); GO_MF:GO:0016788,hydrolase activity, acting on ester bonds# (1e−113); GO_BP:GO:0006629,lipid metabolic process# (1e−113); GO_CC:GO:0005576, extracellularregion# (2e−49) 156 Putative uncharacterized protein n = 2 Tax = Zeamays RepID = B6SQA8_MAIZE (2e−81) 6 56.8 127687297 127688865 157OSJNBa0040D17.12 protein n = 2 Tax = Oryza sativa Japonica Group RepID =Q7XX95_ORYSJ (1e−148); 6 56.8 127714984 127716689 Transferase:Transferase family (1.6e−40); GO_MF:GO:0016747, transferase activity,transferring acyl groups other than amino-acyl groups# (1e−146) 158Putative uncharacterized protein Sb09g005695 n = 3 Tax = AndropogoneaeRepID = C5YU84_SORBI (1e−180); 6 56.8 127766680 127770452GO_MF:GO:0004803, transposase activity# (2e−43); GO_BP:GO:0006313,transposition, DNA-mediated# (2e−43) 159 BZIP transcription factorbZIP109 n = 3 Tax = Glycine max RepID = Q0GPG4_SOYBN (5e−49); DUF1664:Protein 6 56.8 127774370 127787122 of unknown function (DUF1664)(3.1e−67) 160 Putative uncharacterized protein Sb08g000780 n = 1 Tax =Sorghum bicolor RepID = C5YQ53_SORBI (3e−20) 6 56.8 127859677 127860013161 FACT complex subunit SSRP1-B n = 3 Tax = Oryza sativa RepID =SSP1B_ORYSJ (0.0); SSrecog: Structure- 6 56.8 127883928 127890893specific recognition protein (4.6e−144); Rtt106: Histone chaperoneRttp106-like (1.2e−56); HMG_box: HMG (high mobility group) box(3.2e−22); GO_MF:GO:0003677, DNA binding# (0.0); GO_BP:GO:0045449,regulation of transcription# (0.0); GO_CC:GO:0005694, chromosome# (0.0)162 Tetratricopeptide repeat domain protein n = 1 Tax = Microcoleuschthonoplastes PCC 7420 6 56.8 127893148 127895627 RepID = B4VZS1_9CYAN(6e−15); TPR_2: Tetratricopeptide repeat (0.072); TPR_2:Tetratricopeptide repeat (4.7); TPR_2: Tetratricopeptide repeat (28);GO_MF:GO:0005488, binding# (1e−160); GO_BP:GO:0019684, photosynthesis,light reaction# (2e−15); GO_CC:GO:0009941, IDA#chloroplast envelope#(5e−27) 163 Bifunctional protein tilS/hprT n = 3 Tax = AndropogoneaeRepID = B6SSV8_MAIZE (3e−85); Pribosyltran: 6 56.8 127898464 127901020Phosphoribosyl transferase domain (2.3e−24); GO_MF:GO:0016740,transferase activity# (3e−85); GO_BP:GO:0009116, nucleoside metabolicprocess# (3e−85); GO_CC:GO:0005737, cytoplasm# (3e−85) 1643′-N-debenzoyltaxol N-benzoyltransferase-like n = 2 Tax = Oryza sativaRepID = Q9LGF6_ORYSJ (1e−170); 6 56.8 128052937 128055250 Transferase:Transferase family (1.5e−49); GO_MF:GO:0016747, transferase activity,transferring acyl groups other than amino-acyl groups# (0.0) 165 60Sribosomal protein L13 n = 14 Tax = Poaceae RepID = Q7XJB4_ORYSJ (1e−60);Ribosomal_L13e: Ribosomal 6 56.8 128062357 128064852 protein L13e(1.6e−90); GO_MF:GO:0003735, structural constituent of ribosome#(1e−60); GO_BP:GO:0006412, translation# (1e−60); GO_CC:GO:0030529,ribonucleoprotein complex# (1e−60) 166 PRP38 pre-mRNA processing factor38 domain containing B n = 3 Tax = Andropogoneae 6 56.8 128201353128207921 RepID = B6TSE1_MAIZE (1e−112); PRP38: PRP38 family (1.4e−26);DUF1777: Protein of unknown function (DUF1777) (0.022);GO_MF:GO:0004437, inositol or phosphatidylinositol phosphatase activity#(9e−37); GO_BP:GO:0009651, IEP#response to salt stress# (1e−110);GO_CC:GO:0005681, spliceosomal complex# (4e−38) 167 Macrophageerythroblast attacher n = 2 Tax = Andropogoneae RepID = B6TF70_MAIZE(1e−133); 6 56.85 125268872 125272587 GO_MF:GO:0003779, actin binding#(2e−40); GO_BP:GO:0051301, cell division# (2e−40); GO_CC:GO:0016363,nuclear matrix# (2e−40) 168 Protein TOC75, chloroplastic n = 3 Tax =Oryza sativa RepID = TOC75_ORYSJ (2e−22); GO_MF:GO:0015450, 6 56.9125224666 125225259 P—P-bond-hydrolysis-driven protein transmembranetransporter activity# (3e−19); GO_BP:GO:0015031, protein transport#(2e−22); GO_CC:GO:0019867, outer membrane# (2e−22) 169 Ribosomal proteinS27a, isoform CRA_c n = 9 Tax = Euteleostomi RepID = B2RDW1_HUMAN(4e−59); ubiquitin: 6 56.9 125241154 125242015 Ubiquitin family(2.9e−38); Ribosomal_S27: Ribosomal protein S27a (4.2e−30);GO_MF:GO:0003735, structural constituent of ribosome# (1e−57);GO_BP:GO:0006412, translation# (1e−57); GO_CC:GO:0005840, ribosome#(1e−57) 170 Retrotransposon protein, putative, Ty3-gypsy subclass n = 2Tax = Oryza sativa RepID = Q7XGB8_ORYSJ (8e−10); 6 56.9 125241943125247539 GO_MF:GO:0008270, zinc ion binding# (8e−10); GO_BP:GO:0015074,DNA integration# (8e−10) 171 Retrotransposon protein, putative,unclassified n = 1 Tax = Oryza sativa Japonica Group RepID =Q2R3U8_ORYSJ 6 56.9 125247227 125248186 (7e−66); GO_MF:GO:0003677, DNAbinding# (7e−66); GO_BP:GO:0015074, DNA integration# (7e−66) 172WD-repeat protein, putative n = 1 Tax = Ricinus communis RepID =B9RVD2_RICCO (3e−33); 6 56.9 125265027 125267222 GO_BP:GO:0010072,IGI#primary shoot apical meristem specification# (2e−33);GO_CC:GO:0005829, IDA#cytosol# (3e−33) 173 Putative uncharacterizedprotein n = 1 Tax = Zea mays RepID = B6T0R3_MAIZE (4e−13); DVL: DVLfamily 6 56.9 128132458 128133047 (1.1e−08) 174 Protein binding proteinn = 2 Tax = Andropogoneae RepID = B6TV66_MAIZE (0.0); PHD: PHD-finger(0.048); zf- 6 56.9 128159679 128163410 C3HC4: Zinc finger, C3HC4 type(RING finger) (6.8e−09); GO_MF:GO:0046872, metal ion binding# (0.0);GO_CC:GO:0005886, plasma membrane# (5e−23) 175 Putative serine/threonineprotein phosphatase 2A (PP2A) regulatory subunit B′ (B′gamma) n = 1 Tax= Oryza 6 56.9 128299646 128308939 sativa Japonica Group RepID =Q5VRD6_ORYSJ (0.0); B56: Protein phosphatase 2A regulatory B subunit(B56 family) (7.4e−224); GO_MF:GO:0008601, protein phosphatase type 2Aregulator activity# (0.0); GO_BP:GO:0008601, protein phosphatase type 2Aregulator activity# (0.0); GO_CC:GO:0000159, protein phosphatase type 2Acomplex# (0.0) 176 Gibberellin 3-beta-dioxygenase 2-2 n = 4 Tax = Zeamays RepID = B6UAD7_MAIZE (0.0); 2OG-FeII_Oxy: 2OG- 6 56.9 128315121128316850 Fe(II) oxygenase superfamily (3.8e−29); GO_MF:GO:0016702,oxidoreductase activity, acting on single donors with incorporation ofmolecular oxygen, incorporation of two atoms of oxygen# (0.0);GO_BP:GO:0055114, oxidation reduction# (0.0); GO_CC:GO:0005737,cytoplasm# (1e−51) 177 Ankyrin repeat family protein-like n = 2 Tax =Oryza sativa RepID = Q69TB9_ORYSJ (4e−66); Ank: Ankyrin 6 56.9 128318222128321074 repeat (11); Ank: Ankyrin repeat (0.41); Ank: Ankyrin repeat(0.23); Ank: Ankyrin repeat (0.0015); Ank: Ankyrin repeat (2.9); Ank:Ankyrin repeat (0.006); GO_MF:GO:0008234, cysteine-type peptidaseactivity# (4e−66); GO_BP:GO:0006508, proteolysis# (4e−66) 178Cytokinin-O-glucosyltransferase 1 n = 2 Tax = Zea mays RepID =B4FAT6_MAIZE (0.0); UDPGT: UDP- 6 57 128420330 128421841 glucoronosyland UDP-glucosyl transferase (3e−07); GO_MF:GO:0016758, transferaseactivity, transferring hexosyl groups# (0.0); GO_BP:GO:0008152,metabolic process# (0.0); GO_CC:GO:0016021, integral to membrane#(1e−104) 179 Probable cellulose synthase A catalytic subunit 1[UDP-forming] n = 15 Tax = Poaceae RepID = CESA1_ORYSJ 6 57.05 128560617128567283 (0.0); PHD: PHD-finger (0.011); zf-C3HC4: Zinc finger, C3HC4type (RING finger) (0.05); Cellulose_synt: Cellulose synthase (0);GO_MF:GO:0046872, metal ion binding# (0.0); GO_BP:GO:0030244, cellulosebiosynthetic process# (0.0); GO_CC:GO:0016021, integral to membrane#(0.0) 180 Putative uncharacterized protein n = 1 Tax = Zea mays RepID =B6TYR3_MAIZE (2e−39); GO_MF:GO:0005488, 6 57.1 128455019 128455534binding# (9e−11) 181 Putative uncharacterized protein n = 2 Tax = Zeamays RepID = B6SN58_MAIZE (5e−31); GO_MF:GO:0005488, 6 57.1 128465318128466183 binding# (1e−09) 182 Putative uncharacterized proteinSb09g005320 n = 2 Tax = Andropogoneae RepID = C5Z157_SORBI (1e−20) 657.1 128467513 128484308 183 17.5 kDa class II heat shock protein n = 1Tax = Zea mays RepID = B6U175_MAIZE (3e−52); HSP20: 6 57.1 128470652128471159 Hsp20/alpha crystallin family (0.0014); GO_MF:GO:0051082,unfolded protein binding# (9e−11); GO_BP:GO:0006950, response to stress#(3e−52); GO_CC:GO:0005737, cytoplasm# (1e−35) 184 Putativeuncharacterized protein Sb04g007000 n = 1 Tax = Sorghum bicolor RepID =C5XXW4_SORBI (2e−10) 6 57.1 128501719 128502463 185 Putativewall-associated serine/threonine kinase n = 1 Tax = Oryza sativaJaponica Group 6 57.1 128520533 128525078 RepID = Q6ZK05_ORYSJ (1e−177);Pkinase: Protein kinase domain (2.2e−39); Pkinase_Tyr: Protein tyrosinekinase (3.6e−34); GO_MF:GO:0016301, kinase activity# (1e−177);GO_BP:GO:0016301, kinase activity# (1e−177) 186 Lactoylglutathione lyasen = 1 Tax = Zea mays RepID = B6UGW8_MAIZE (2e−92); Glyoxalase: 6 57.1128580829 128583989 Glyoxalase/Bleomycin resistance protein/Dioxygenasesuperfamily (7e−07); GO_MF:GO:0016829, lyase activity# (2e−92) 187Isoform 2 of Probable protein phosphatase 2C 48 n = 1 Tax = Oryza sativaJaponica Group RepID = Q6L482-2 6 57.2 128601099 128602653 (3e−10);GO_MF:GO:0046872, metal ion binding# (4e−10); GO_BP:GO:0004721,phosphoprotein phosphatase activity# (4e−10) 188 Transposon protein,putative, CACTA, En/Spm sub-class n = 1 Tax = Oryza sativa JaponicaGroup 6 57.2 128635663 128637800 RepID = Q10N80_ORYSJ (2e−50);GO_MF:GO:0004803, transposase activity# (2e−50); GO_BP:GO:0006313,transposition, DNA-mediated# (2e−50); GO_CC:GO:0005783, IDA#endoplasmicreticulum# (2e−27) 189 Nucleotide sugar translocator BT2A n = 4 Tax =Zea mays RepID = B2LWG5_MAIZE (0.0); Mito_carr: 6 57.2 128638388128642377 Mitochondrial carrier protein (6.6e−26); Mito_carr:Mitochondrial carrier protein (1.1e−34); Mito_carr: Mitochondrialcarrier protein (5.9e−33); GO_MF:GO:0005488, binding# (0.0);GO_BP:GO:0055085, transmembrane transport# (0.0); GO_CC:GO:0016021,integral to membrane# (0.0) 190 Putative uncharacterized protein n = 1Tax = Zea mays RepID = B6SUB9_MAIZE (4e−99) 6 57.3 128663003 128666303191 OSJNBa0053K19.6 protein n = 5 Tax = Poaceae RepID = Q7X809_ORYSJ(1e−58); GO_MF:GO:0016491, 6 57.3 128667645 128669160 oxidoreductaseactivity# (1e−58); GO_BP:GO:0055114, oxidation reduction# (1e−58);GO_CC:GO:0005777, IDA#peroxisome# (1e−44) 192 Phospholipase D alpha 1 n= 7 Tax = Poaceae RepID = PLDA1_ORYSJ (0.0); C2: C2 domain (7.6e−06);PLDc: 6 57.4 128722429 128725039 Phospholipase D. Active site motif(1.2e−09); PLDc: Phospholipase D. Active site motif (1.4e−08);GO_MF:GO:0005509, calcium ion storage activity# (0.0); GO_BP:GO:0046470,phosphatidylcholine metabolic process# (0.0); GO_CC:GO:0016020,membrane# (0.0) 193 Putative uncharacterized protein n = 1 Tax = Zeamays RepID = B4FWV4_MAIZE (4e−69) 6 57.4 128734831 128736354 194CDPK-related protein kinase n = 1 Tax = Zea mays RepID = B6SYP7_MAIZE(2e−67); Pkinase: Protein kinase 6 57.5 128754439 128759348 domain(3.2e−09); Pkinase_Tyr: Protein tyrosine kinase (3.5e−06); WD40: WDdomain, G-beta repeat (0.011); WD40: WD domain, G-beta repeat (0.0046);GO_MF:GO:0016740, transferase activity# (9e−87); GO_BP:GO:0016301,kinase activity# (9e−87); GO_CC:GO:0005886, plasma membrane# (1e−55) 195Putative uncharacterized protein n = 2 Tax = Zea mays RepID =Q5GAU8_MAIZE (9e−27) 6 57.5 128781974 128782353 196 NBS-LRR classdisease resistance protein n = 1 Tax = Oryza sativa Japonica Group RepID= B5UBC0_ORYSJ 6 57.5 128790205 128798880 (0.0); NB-ARC: NB-ARC domain(1.1e−35); GO_MF:GO:0017111, nucleoside-triphosphatase activity# (0.0);GO_BP:GO:0006952, defense response# (0.0) 197 Putative uncharacterizedprotein n = 2 Tax = Zea mays RepID = B6TX07_MAIZE (1e−112); IQ: IQcalmodulin- 6 57.6 128819708 128821261 binding motif (7.9e−05); IQ: IQcalmodulin-binding motif (0.0012) 198 Putative Mlal n = 1 Tax = Sorghumbicolor RepID = Q8LJZ8_SORBI (1e−143); NB-ARC: NB-ARC domain 6 57.6128825273 128833947 (1.4e−57); NACHT: NACHT domain (0.036); LRR_1:Leucine Rich Repeat (9); LRR_1: Leucine Rich Repeat (2.4); LRR_1:Leucine Rich Repeat (1.6); LRR_1: Leucine Rich Repeat (45); LRR_1:Leucine Rich Repeat (3e+02); LRR_1: Leucine Rich Repeat (43);GO_MF:GO:0005524, ATP binding# (1e−149); GO_BP:GO:0006952, defenseresponse# (1e−149) 199 Putative RGH1A n = 1 Tax = Oryza sativa JaponicaGroup RepID = Q6Z021_ORYSJ (1e−16); 6 57.6 128827662 128828455GO_MF:GO:0005524, ATP binding# (1e−16); GO_BP:GO:0006952, defenseresponse# (1e−16) 200 Putative serine/threonine protein kinase(Fragment) n = 1 Tax = Oryza sativa Japonica Group 6 57.6 128927057128927892 RepID = Q84P73_ORYSJ (9e−16); GO_MF:GO:0005524, ATP binding#(4e−17); GO_BP:GO:0016567, IGI#protein ubiquitination# (4e−17);GO_CC:GO:0000151, ubiquitin ligase complex# (4e−17) 201 PutativeAvr9/Cf-9 rapidly elicited protein n = 1 Tax = Oryza sativa JaponicaGroup RepID = 6 57.6 129025487 129026094 Q6EUK7_ORYSJ (1e−09);GO_MF:GO:0005488, binding# (1e−09); GO_BP:GO:0016567, IGI#proteinubiquitination# (1e−09); GO_CC:GO:0000151, ubiquitin ligase complex#(1e−09) 202 H0716A07.11 protein n = 1 Tax = Oryza sativa RepID =Q01MA7_ORYSA (0.0); Inhibitor_I9: Peptidase inhibitor 6 57.6 129031227129036927 I9 (1.2e−19); Peptidase_S8: Subtilase family (3.6e−10); PA: PAdomain (9.8e−05); GO_MF:GO:0043086, negative regulation of catalyticactivity# (0.0); GO_BP:GO:0043086, negative regulation of catalyticactivity# (0.0); GO_CC:GO:0009505, IDA#expansin# (1e−153) 203Inositol-3-phosphate synthase n = 16 Tax = Magnoliophyta RepID =INO1_ORYSJ (1e−27); GO_MF:GO:0016853, 6 57.6 129042421 129042715isomerase activity# (1e−27); GO_BP:GO:0008654, phospholipid biosyntheticprocess# (1e−27); GO_CC:GO:0005737, cytoplasm# (1e−27) 204Inositol-3-phosphate synthase n = 16 Tax = Magnoliophyta RepID =INO1_ORYSJ (1e−108); NAD_binding_5: 6 57.6 129042809 129044242Myo-inositol-1-phosphate synthase (3.4e−08); Inos-1-P_synth:Myo-inositol-1-phosphate synthase (2.2e−61); GO_MF:GO:0016853, isomeraseactivity# (1e−108); GO_BP:GO:0008654, phospholipid biosynthetic process#(1e−108); GO_CC:GO:0005737, cytoplasm# (1e−108) 205 Leaf senescencerelated protein-like n = 2 Tax = Oryza sativa RepID = Q69RQ8_ORYSJ(2e−18); IBB: Importin 6 57.6 129045552 129049517 beta binding domain(0.076); GO_MF:GO:0008565, protein transporter activity# (1e−14);GO_BP:GO:0015031, protein transport# (1e−14); GO_CC:GO:0048471,ISS#perinuclear region of cytoplasm# (1e−14) 206 Xylanase inhibitorprotein 1 n = 1 Tax = Zea mays RepID = B6U2X8_MAIZE (1e−170);Glyco_hydro_18: Glycosyl 6 57.6 129086009 129087210 hydrolases family 18(1.2e−19); GO_MF:GO:0043169, cation binding# (1e−170); GO_BP:GO:0045493,xylan catabolic process# (1e−170); GO_CC:GO:0005576, extracellularregion# (2e−87) 207 Putative uncharacterized protein n = 1 Tax = Oryzasativa Indica Group RepID = A2YUK8_ORYSI (1e−15) 6 57.6 129126147129126488 208 Putative transposon protein n = 1 Tax = Oryza sativaJaponica Group RepID = Q8H801_ORYSJ (3e−31); 6 57.6 129127693 129129065Transposase_23: TNP1/EN/SPM transposase (6.2e−05) 209 Putativeuncharacterized protein Sb05g026840 n = 1 Tax = Sorghum bicolor RepID =C5Y811_SORBI (0.0); 6 57.6 129138400 129140529 DUF594: Protein ofunknown function, DUF594 (1.6e−30); GO_MF:GO:0046872, metal ion binding#(1e−115); GO_BP:GO:0006278, RNA-dependent DNA replication# (1e−51) 210Retrotransposon protein, putative, unclassified n = 1 Tax = Oryza sativaJaponica Group RepID = Q2QQR5_ORYSJ 6 57.6 129142269 129144473 (3e−33);zf-CCHC: Zinc knuckle (0.015); GO_MF:GO:0008270, zinc ion binding#(2e−49); GO_BP:GO:0006278, RNA-dependent DNA replication# (3e−40) 211HAT family dimerisation domain containing protein n = 1 Tax = Oryzasativa Japonica Group 6 57.7 129169527 129171992 RepID = Q2R0F0_ORYSJ(1e−107); zf-BED: BED zinc finger (1.4e−09); hATC: hAT familydimerisation domain (1.5e−28); GO_MF:GO:0046983, protein dimerizationactivity# (1e−116); GO_BP:GO:0006350, transcription# (1e−33) 212 Triosephosphate/phosphate translocator, non-green plastid, chloroplast,putative n = 1 Tax = Ricinus communis 6 57.7 129222041 129225144 RepID =B9RB11_RICCO (1e−116); UAA: UAA transporter family (0.0016); DUF6:Integral membrane protein DUF6 (1.2e−11); DUF6: Integral membraneprotein DUF6 (0.0068); TPT: Triose-phosphate Transporter family(3.5e−52); GO_MF:GO:0005215, transporter activity# (1e−146);GO_BP:GO:0006810, transport# (1e−146); GO_CC:GO:0016021, integral tomembrane# (1e−146) 213 Putative transcription regulatory protein n = 2Tax = Oryza sativa RepID = Q94LQ9_ORYSJ (2e−68); U-box: 6 57.7 129243446129247172 U-box domain (0.0089); GO_MF:GO:0016301, kinase activity#(2e−79); GO_BP:GO:0016301, kinase activity# (2e−79); GO_CC:GO:0000151,ubiquitin ligase complex# (3e−25) 214 NADH-ubiquinone oxidoreductase10.5 kDa subunit n = 4 Tax = Andropogoneae RepID = B6TDN0_MAIZE 6 57.8129361190 129365442 (2e−31); L51_S25_CI-B8: Mitochondrial ribosomalprotein L51/S25/CI-B8 domain (2e−15); GO_MF:GO:0016491, oxidoreductaseactivity# (4e−18); GO_BP:GO:0055114, oxidation reduction# (4e−18);GO_CC:GO:0045271, IDA#respiratory chain complex I# (4e−21) 215 Putativeuncharacterized protein n = 1 Tax = Zea mays RepID = B6TDS4_MAIZE(1e−09) 6 57.9 129437152 129437697 216 Putative receptor protein kinasen = 1 Tax = Oryza sativa Japonica Group RepID = Q65XS7_ORYSJ (0.0); 657.95 129472709 129517525 LRRNT_2: Leucine rich repeat N-terminal domain(6.8e−13); LRR_1: Leucine Rich Repeat (3.2); LRR_1: Leucine Rich Repeat(0.91); LRR_1: Leucine Rich Repeat (0.11); LRR_1: Leucine Rich Repeat(5.8); LRR_1: Leucine Rich Repeat (12); LRR_1: Leucine Rich Repeat (41);LRR_1: Leucine Rich Repeat (1.2); LRR_1: Leucine Rich Repeat (4.3);LRR_1: Leucine Rich Repeat (0.09); LRR_1: Leucine Rich Repeat (2.5);LRR_1: Leucine Rich Repeat (0.11); LRR_1: Leucine Rich Repeat (1.2);LRR_1: Leucine Rich Repeat (0.21); LRR_1: Leucine Rich Repeat (2.3);LRR_1: Leucine Rich Repeat (2.8); LRR_1: Leucine Rich Repeat (0.13);LRR_1: Leucine Rich Repeat (0.1); LRR_1: Leucine Rich Repeat (0.29);LRR_1: Leucine Rich Repeat (0.13); LRR_1: Leucine Rich Repeat (0.44);LRR_1: Leucine Rich Repeat (1.7); LRR_1: Leucine Rich Repeat (0.041);LRR_1: Leucine Rich Repeat (0.26); LRR_1: Leucine Rich Repeat (0.21);LRR_1: Leucine Rich Repeat (0.96); LRR_1: Leucine Rich Repeat (14);LRR_1: Leucine Rich Repeat (0.94); LRR_1: Leucine Rich Repeat (0.092);LRR_1: Leucine Rich Repeat (0.25); LRR_1: Leucine Rich Repeat (5.5);LRR_1: Leucine Rich Repeat (6); LRR_1: Leucine Rich Repeat (12); LRR_1:Leucine Rich Repeat (0.33); LRR_1: Leucine Rich Repeat (5.1); LRR_1:Leucine Rich Repeat (0.078); LRR_1: Leucine Rich Repeat (2.2); LRR_1:Leucine Rich Repeat (3.4); LRR_1: Leucine Rich Repeat (0.56); LRR_1:Leucine Rich Repeat (1.2); LRR_1: Leucine Rich Repeat (3.3); LRR_1:Leucine Rich Repeat (2.7); LRR_1: Leucine Rich Repeat (47); LRR_1:Leucine Rich Repeat (0.85); GO_MF:GO:0016301, kinase activity# (0.0);GO_BP:GO:0016301, kinase activity# (0.0); GO_CC:GO:0016021, integral tomembrane# (0.0) 217 BEL1-related homeotic protein 30 n = 2 Tax =Andropogoneae RepID = B6SWM4_MAIZE (1e−64); 6 58 129501726 129502856GO_MF:GO:0043565, sequence-specific DNA binding# (1e−64);GO_BP:GO:0045449, regulation of transcription# (1e−64);GO_CC:GO:0005634, nucleus# (1e−64) 218 HAT family dimerization domainprotein n = 2 Tax = Oryza sativa RepID = D0UZH7_ORYSJ (6e−94); 6 58129520475 129521414 GO_MF:GO:0046983, protein dimerization activity#(3e−86) 219 Putative uncharacterized protein n = 1 Tax = Zea mays RepID= C0PP88_MAIZE (1e−46) 6 58 129538411 129540061 220 Putativeuncharacterized protein Sb01g027800 n = 1 Tax = Sorghum bicolor RepID =C5WQN1_SORBI (3e−13) 6 58 129581574 129582554 221 Alliin lyase n = 3 Tax= Zea mays RepID = B6TK37_MAIZE (0.0); Aminotran_1_2: Aminotransferaseclass I and II 6 58 129661095 129664534 (0.006); Alliinase_C: Allinase,C-terminal domain (4.8e−199); GO_MF:GO:0030170, pyridoxal phosphatebinding# (0.0); GO_BP:GO:0080022, LMP#primary root development# (9e−82);GO_CC:GO:0005737, cytoplasm# (9e−82) 222 O-sialoglycoproteinendopeptidase, putative n = 2 Tax = rosids RepID = B9T542_RICCO(1e−164); 6 58 129670272 129674945 Peptidase_M22: Glycoprotease family(6.3e−73); GO_MF:GO:0008270, zinc ion binding# (0.0); GO_BP:GO:0006508,proteolysis# (0.0); GO_CC:GO:0005737, cytoplasm# (1e−125) 223 HAT familydimerisation domain containing protein n = 2 Tax = Oryza sativa JaponicaGroup 6 58 129679558 129681972 RepID = Q7XE06_ORYSJ (1e−44); hATC: hATfamily dimerisation domain (1e−11); GO_MF:GO:0046983, proteindimerization activity# (7e−52); GO_BP:GO:0006350, transcription# (7e−52)224 Nonspecific lipid-transfer protein n = 2 Tax = Zea mays RepID =B6SZZ6_MAIZE (6e−29); Tryp_alpha_amyl: 6 58 129703134 129703976 Proteaseinhibitor/seed storage/LTP f (8.5e−10); GO_MF:GO:0008289, lipid binding#(3e−13); GO_BP:GO:0006869, lipid transport# (6e−29) 225 Stem 28 kDaglycoprotein n = 1 Tax = Zea mays RepID = B6T003_MAIZE (1e−147);Acid_phosphat_B: HAD 6 58.2 129792745 129795009 superfamily, subfamilyIIIB (Acid (3.6e−90); GO_MF:GO:0003993, acid phosphatase activity#(1e−147); GO_BP:GO:0003993, acid phosphatase activity# (1e−147);GO_CC:GO:0005886, plasma membrane# (1e−51) 226 Serine-threonine proteinkinase, plant-type, putative n = 1 Tax = Ricinus communis RepID =B9RKF0_RICCO 6 58.3 129841439 129842719 (1e−105); LRRNT_2: Leucine richrepeat N-terminal domain (2.6e−06); LRR_1: Leucine Rich Repeat (3.9);LRR_1: Leucine Rich Repeat (1.8); LRR_1: Leucine Rich Repeat (5.4);LRR_1: Leucine Rich Repeat (32); LRR_1: Leucine Rich Repeat (2.1e+02);LRR_1: Leucine Rich Repeat (70); LRR_1: Leucine Rich Repeat (0.72);LRR_1: Leucine Rich Repeat (0.23); LRR_1: Leucine Rich Repeat (1.4e+02);GO_MF:GO:0005515, protein binding# (1e−141); GO_BP:GO:0055114, oxidationreduction# (1e−105); GO_CC:GO:0009505, IDA#expansin# (2e−68) 227Putative uncharacterized protein n = 4 Tax = Zea mays RepID =B6U1N7_MAIZE (7e−24) 6 58.3 129880459 129880719 228 Retrotransposonprotein, putative, Ty1-copia subclass n = 2 Tax = Oryza sativa RepID =Q7XH58_ORYSJ (6e−28); 6 58.3 129887372 129887779 GO_MF:GO:0003677, DNAbinding# (6e−28); GO_BP:GO:0015074, DNA integration# (6e−28) 229OSJNBa0064G10.18 protein n = 3 Tax = Oryza sativa RepID = Q7XKB3_ORYSJ(7e−94); WD40: WD domain, G- 6 58.35 129888300 129891431 beta repeat(1.6); WD40: WD domain, G-beta repeat (0.016); WD40: WD domain, G-betarepeat (3e−09); WD40: WD domain, G-beta repeat (3.4e−12); WD40: WDdomain, G-beta repeat (2.6e−10); GO_MF:GO:0016740, transferase activity#(5e−92); GO_BP:GO:0016905, myosin heavy chain kinase activity# (2e−84);GO_CC:GO:0005874, microtubule# (1e−89) 230 SIN3 component, histonedeacetylase complex n = 1 Tax = Populus trichocarpa RepID = B9HU88_POPTR(2e−15); 6 58.4 129892633 129892893 GO_MF:GO:0016564, transcriptionrepressor activity# (1e−12); GO_BP:GO:0006355, regulation oftranscription, DNA-dependent# (5e−21); GO_CC:GO:0005634, nucleus#(5e−21) 231 L-lactate dehydrogenase n = 3 Tax = Oryza sativa RepID =Q0E4Q5_ORYSJ (1e−51); Ldh_1_C: lactate/malate 6 58.4 129892886 129893304dehydrogenase, alpha/b (7.2e−05); GO_MF:GO:0016616, oxidoreductaseactivity, acting on the CH—OH group of donors, NAD or NADP as acceptor#(1e−51); GO_BP:GO:0055114, oxidation reduction# (1e−51);GO_CC:GO:0005737, cytoplasm# (1e−51) 232 Derlin-1.2 n = 2 Tax = Zea maysRepID = DER12_MAIZE (1e−137); DER1: Der1-like family (2.5e−56); 6 58.4129897727 129902466 GO_MF:GO:0005515, protein binding# (3e−35);GO_BP:GO:0006950, response to stress# (1e−137); GO_CC:GO:0016021,integral to membrane# (1e−137) 233 Putative uncharacterized protein n =1 Tax = Zea mays RepID = B6UF52_MAIZE (1e−14) 6 58.5 129955339 129955661234 OSJNBa0036E02.10 protein n = 2 Tax = Oryza sativa RepID =Q7F7E1_ORYSJ (7e−35); IQ: IQ calmodulin- 6 58.5 129963265 129965399binding motif (0.0013); IQ: IQ calmodulin-binding motif (0.006) 235Hexokinase-1 n = 2 Tax = Zea mays RepID = B6TL75_MAIZE (0.0);Hexokinase_1: Hexokinase (2.9e−09); 6 58.5 129979002 129982104Hexokinase_2: Hexokinase (4e−79); GO_MF:GO:0016773, phosphotransferaseactivity, alcohol group as acceptor# (0.0); GO_BP:GO:0016301, kinaseactivity# (0.0); GO_CC:GO:0005737, cytoplasm# (1e−152) 236Retrotransposon protein, putative, unclassified n = 1 Tax = Oryza sativaJaponica Group RepID = Q7XE51_ORYSJ 6 58.5 129986022 129989105 (7e−30);zf-CCHC: Zinc knuckle (0.0013); zf-CCHC: Zinc knuckle (0.16);GO_MF:GO:0008270, zinc ion binding# (7e−44); GO_BP:GO:0006278,RNA-dependent DNA replication# (4e−35) 237 IAA16-auxin-responsiveAux/IAA family member n = 2 Tax = Zea mays RepID = B6TT61_MAIZE (5e−89);6 58.6 130004376 130008214 AUX_IAA: AUX/IAA family (2.4e−19);GO_MF:GO:0046983, protein dimerization activity# (5e−89);GO_BP:GO:0045449, regulation of transcription# (5e−89);GO_CC:GO:0005634, nucleus# (5e−89) 238 Os07g0695700 protein n = 1 Tax =Oryza sativa Japonica Group RepID = Q0D3B3_ORYSJ (1e−12) 6 58.6130011779 130011936 239 Histidine-containing phosphotransfer protein 4 n= 2 Tax = Andropogoneae RepID = B6SRE6_MAIZE (2e−50); 6 58.6 130159712130161359 Hpt: Hpt domain (1.7e−08); GO_MF:GO:0004871, signal transduceractivity# (7e−65); GO_BP:GO:0004871, signal transducer activity#(7e−65); GO_CC:GO:0005737, cytoplasm# (2e−38) 240 OSJNBa0027O01.13protein n = 3 Tax = Oryza sativa RepID = Q7XXC5_ORYSJ (0.0); NCD3G: NineCysteines 6 58.6 130239488 130245375 Domain of family 3 GPC (0.093);GO_CC:GO:0005773, IDA#vacuole# (0.0) 241 Putative polyprotein n = 1 Tax= Zea mays RepID = Q8SA93_MAIZE (3e−59); GO_MF:GO:0003964, RNA- 6 58.6130257021 130258701 directed DNA polymerase, group II intron encoded#(3e−59); GO_BP:GO:0015074, DNA integration# (3e−59); GO_CC:GO:0005634,nucleus# (3e−59) † cM = centiMorgans. †† bp = base pair of ArizonaGenomics Institute B73 RefGen_v2 sequence.

As various modifications could be made in the constructions and methodsherein described and illustrated without departing from the scope of theinvention, it is intended that all matter contained in the foregoingdescription or shown in the accompanying drawings shall be interpretedas illustrative rather than limiting. The breadth and scope of thepresent invention should not be limited by any of the above-describedexemplary embodiments, but should be defined only in accordance with thefollowing claims appended hereto and their equivalents. All patent andnon-patent documents cited in this specification are incorporated hereinby reference in their entireties.

1. A method of obtaining a corn plant with enhanced anthracnose stalkrot resistance, said method comprising: a) obtaining a DNA sample fromat least one corn plant or part thereof from a population of cornplants; b) detecting in the DNA sample the presence of an anthracnosestalk rot resistance allele, wherein said allele is within 5 cM of a “A”corresponding to position 151 of SEQ ID NO:106, and wherein the “A” atsaid position is associated with enhanced anthracnose stalk rotresistance; c) selecting at least a first plant comprising said alleleand enhanced anthracnose stalk rot resistance compared to a plantlacking said allele; d) crossing the plant selected in step c) with asecond corn plant, wherein said second corn plant lacks or isheterozygous for said anthracnose stalk rot resistance allele; e)collecting seeds from the cross of step d); and f) growing at least oneprogeny corn plant from the seeds of step e); wherein said progeny cornplant comprises said allele and has enhanced anthracnose stalk rotresistance compared to a corn plant lacking said allele.
 2. The methodof claim 1, wherein said selecting comprises detecting a polymorphismlocated in a chromosomal segment flanked by marker loci AY107053 andumc1379.
 3. The method of claim 2, wherein said polymorphism is locatedin a chromosomal segment flanked by marker loci SEQ ID NO: 4 or 52 andSEQ ID NO:
 5. 4. The method of claim 2, wherein said polymorphism islocated in a chromosomal segment flanked by marker loci SEQ ID NO: 81and SEQ ID NO: 2 or
 55. 5. The method of claim 4, wherein saidchromosomal segment is flanked by marker loci SEQ ID NO: 81 and SEQ IDNO:
 54. 6. The method of claim 5, wherein said chromosomal segment isflanked by marker loci SEQ ID NO: 82 and SEQ ID NO:
 54. 7-32. (canceled)33. The method of claim 1, wherein the selecting comprises detectingenhanced anthracnose stalk rot resistance, wherein the detectingenhanced anthracnose stalk rot resistance comprises detecting apolynucleotide comprising SEQ ID NO: 106 in the DNA sample.